The indices of antipneumococcal immunity in children with chronic pneumonia

In 76 children with chronic pneumonia the levels of serum antibodies to pneumococcal capsular polysaccharide antigens (serotypes 1, 3, 6B, 8, 9N, 15F, 23F), O-polysaccharide and pneumococcal protein somatic antigens were determined by ELISA techniques. The study showed that in sick children the content of antipneumococcal antibodies in the blood increased with age. No correlation between the content of total immunoglobulins and that of antipneumococcal antibodies in the blood of the patients was established, but a sharp decrease in the concentration of antibodies was registered in a child with hypoglobulinemia. No increase in the level of antibodies to pneumococcal antigens was observed in cases of the exacerbation of Pneumococcus-induced inflammatory process in the lungs.     

Epidemiologic problems of pneumococcal infection

During the microbiological examination of 314 tentatively healthy donors, residents of Leningrad, Streptococcus pneumoniae was detected in the rhinopharynx of 94 donors (29.9%) in amounts ranging from a few individual microbial cells to 10(6) microbial cells with the mean geometric concentration equal to 3.58 lg. Antipneumococcal antibodies were detected in the blood of 147 donors (46.8%). The occurrence of antipneumococcal antibodies was approximately 2.5 times higher than the average level of carriership, but in most cases the dynamics of immunological reactions changed in the same direction. Considerable monthly and annual fluctuations in the occurrence of pneumococcal carriership and antipneumococcal antibodies, as well as their correlation with the epidemic rises of acute respiratory diseases, influenza and other acute respiratory viral diseases, were established. These data indicate that pneumococcal infection plays a definite role in the epidemic rises of acute respiratory diseases.     

Antibodies to Streptococcus pneumoniae antigens in newborn infants

The levels of antibodies to capsular polysaccharide antigens of pneumococci (serotypes 1, 3, 6B, 8, 9N, 15F, 23F), C-polysaccharide and protein antigen of pneumococci in the blood sera of 38 newborn infants at the moment of their birth (umbilical blood) and on the 5th or 6th day of their life, in their mothers' blood sera, as well as in the colostrum and milk of 48 nursing women, have been studied by means of the enzyme immunoassay. The study showed that in the normal course of pregnancy antibodies to pneumococci were transferred transplacentally from the mother to the fetus. Though in most cases their content in the blood of newborn infants was lower than that in maternal blood, it exceeded the average level of antipneumococcal antibodies in children aged 3-12 months. In the milk of nursing mothers considerable amount of IgA antibodies to pneumococci was detected, which might be an additional protective factor with respect to pneumococcal infection in infants.     

Population pattern of pneumococci with lower susceptibility to penicillin and prospects of antipneumococcal vaccination to control antibiotic resistance distribution

Large-scale antipneumococcal vaccination is followed by changes in the serotype composition and level of antibiotic resistance in pneumococci. The aim of the study was to evaluate the serotype composition and population pattern of pneumococci with lower susceptibility to penicillin before large-scale antipneumococcal vaccination. Among 260 Streptococcus pneumoniae strains isolated in the Russian Federation within 2003-2007, serotypes 23F (37.2%) and 19F (13.9%) were the most frequent ones. 19.3% of the isolates belonged to serogroup 6, 3.6% of the isolates each belonged to serotype 3 and serogroup 18, 4.9% of the isolates belonged to serotype 14 and 2.2% of the isolates belonged to serotype 19A. 66.8% of the isolates belonged to serotypes of the 7-valent conjugated pneumococcal vaccine, 67.3 and 82.1% of the isolates belonged to the 10- and 13-valent conjugated pneumococcal vaccines respectively. The isolates with lower susceptibility to penicillin were characterized by significant clonality and 56.9% of them belonged to 4 global clonal complexes (CC81, CC156, CC320 and CC315). Inclusion of the conjugated antipneumococcal vaccine to the National Vaccination Time-Table of the Russian Federation could promote lower levels of antibiotic resistance in pneumococci.     

Impaired antipneumococcal antibody production in patients without spleens.

Fifteen splenectomised and 15 normal subjects were studied, in absence of any intentional immunisation, for pokeweed mitogen induced synthesis of antipneumococcal capsular polysaccharide antibodies in vitro by peripheral blood mononuclear cells. Results showed that removal of the spleen had caused a persistent immune deficiency of circulating B cells capable of synthesising IgM antipneumococcal capsular polysaccharide. In vitro synthesis of polyclonal IgM and IgG by peripheral blood mononuclear cells of subjects without spleens was also depressed. These defects were due to an abnormality of the B cell compartment. These data are evidence of the major role of the spleen in the control and production of a consistent part of pokeweed mitogen responsive circulating B cells and add another facet to the complex immune dysfunction of splenectomised subjects. The findings, moreover, may help in understanding the susceptibility of splenectomised people to pneumococcal sepsis and the delayed and impaired antibody response to pneumococcal vaccine.     

Cross-reactivity of the membrane teichoic acid of Streptococcus faecalis N.C.I.B. 8191 in antipneumococcal type XII and type XVI sera

The membrane teichoic acid (MTA) precipitates heavily in the equine and rabbit antipneumococcal (anti-Pn) type XII sera tested. MTA contains multiple kojibiosyl residues and this reaction confirms the presence of kojibiosyl residues in the capsular polysaccharide, S XII, the antigenic determinant of pneumococcal type XII. MTA also precipitates one-third of the antibodies in an anti-Pn XVI serum: the poly(glycerol phosphate) residues of MTA may be involved in this reaction as well as d-glucose residues. Precipitation of MTA in anti-Pn XII is strongly inhibited by kojibiose, but the reaction in anti-Pn XVI is unaffected.     

Association of pneumococcal carriage in infants with the risk of carriage among their contacts in Nha Trang,Vietnam: A nested cross-sectional survey

BackgroundInfants are at highest risk of pneumococcal disease. Their added protection through herd effects is a key part in the considerations on optimal pneumococcal vaccination strategies. Yet, little is currently known about the main transmission pathways to this vulnerable age group. Hence, this study investigates pneumococcal transmission routes to infants in the coastal city of Nha Trang, Vietnam.Methods and findingsIn October 2018, we conducted a nested cross-sectional contact and pneumococcal carriage survey in randomly selected 4- to 11-month-old infants across all 27 communes of Nha Trang. Bayesian logistic regression models were used to estimate age specific carriage prevalence in the population, a proxy for the probability that a contact of a given age could lead to pneumococcal exposure for the infant. We used another Bayesian logistic regression model to estimate the correlation between infant carriage and the probability that at least one of their reported contacts carried pneumococci, controlling for age and locality. In total, 1,583 infants between 4 and 13 months old participated, with 7,428 contacts reported. Few infants (5%, or 86 infants) attended day care, and carriage prevalence was 22% (353 infants). Most infants (61%, or 966 infants) had less than a 25% probability to have had close contact with a pneumococcal carrier on the surveyed day. Pneumococcal infection risk and contact behaviour were highly correlated: If adjusted for age and locality, the odds of an infant’s carriage increased by 22% (95% confidence interval (CI): 15 to 29) per 10 percentage points increase in the probability to have had close contact with at least 1 pneumococcal carrier. Moreover, 2- to 6-year-old children contributed 51% (95% CI: 39 to 63) to the total direct pneumococcal exposure risks to infants in this setting. The main limitation of this study is that exposure risk was assessed indirectly by the age-dependent propensity for carriage of a contact and not by assessing carriage of such contacts directly.ConclusionsIn this study, we observed that cross-sectional contact and infection studies could help identify pneumococcal transmission routes and that preschool-age children may be the largest reservoir for pneumococcal transmission to infants in Nha Trang, Vietnam.     

Structural and functional studies in vitro on the p6 protein from the HIV-1 gag open reading frame

Protein p6 from HIV-1 gag open reading frame is reported to affect both the final phase of assembly of the viral particle and the early stage of the gag polyprotein maturation in vitro. Two separate hypotheses have been proposed, on only one of these reported effects. We think that both observations may be eventually explained if p6 protein strongly inhibits the HIV-1 proteinase. Protein p6 was synthesised by solid-phase peptide synthesis. Several methods of folding the p6 protein were tested, each resulting in the random structure according to both CD and 1D proton NMR spectra. A uniformly high exposure of NH protons to the solution was confirmed by temperature-dependent NMR spectra and isotope exchange experiments. Thus the p6 protein does not have any rigid conformation in solution. A rigid structure is not formed after further cleavage by HIV-1 proteinase as neither the protein nor its fragments are cleaved by this proteinase. In addition, the p6 protein itself does not act as inhibitor of HIV-1 proteinase. This excludes a direct role of p6 protein and supports the hypothesis that p6 is involved in forming the appropriate structure of gag polyprotein precursor. The role of slowly cleaved tight gag-proteinase in the final stage of maturation may be to slow down maturation of the precursor polyproteins prior to their transport to final location in the membrane.     

Pneumococcal antibody concentrations of subjects in communities fully or partially vaccinated with a seven-valent pneumococcal conjugate vaccine

Background

A recent trial with PCV-7 in a rural Gambian community showed reduced vaccine-type pneumococcal carriage in fully vaccinated compared with control communities. We measured pneumococcal polysaccharide antibody concentrations in this trial to understand further the mechanisms underlying the observed changes.

Methods

A single-blind, cluster-randomized (by village) trial was conducted in 21 Gambian villages. In 11 villages, all residents received PCV-7 (Vaccine group); in 10 control villages only children <30 months old or those born during the study received PCV-7. Subjects over the age of 30 months resident in vaccine villages received a single dose of PCV-7 whilst those in control villages received a single dose of a serogroup C meningococcal conjugate vaccine. Serum antibody concentrations against specific pneumococcal polysaccharides were measured in approximately 200 age-stratified subjects before, 4–6, 12 and 24 months following vaccination.

Results

Baseline pneumococcal antibody concentrations were generally high and increased with age up to 10 years. One dose of PCV-7 increased geometric mean antibody concentrations (GMC) in vaccinated versus control villages for vaccine serotypes 6B and 18C, and 4 and 18C, in the young (under 5 years) and older age groups (5+ years) respectively. There were significantly higher proportions of subjects in the vaccinated than in the control communities with an antibody concentration believed to protect against carriage (>5.0 µg/mL) for all but serotype 9V of the PCV-7 serotypes in the older group, but not in the younger age group.

Conclusion

Higher antibodies in vaccinated communities provide an explanation for the lower pneumococcal carriage rates in fully vaccinated compared to control communities.

Trial Registration

Controlled-Trials.com ISRCTN51695599 51695599.     

Lactococcal proteinase maturation protein PrtM is a lipoprotein.

The production of enzymatically active proteinase by lactococci requires the joint presence of a proteinase gene, prtP, and a gene encoding a maturation protein, prtM. A 32-kDa protein produced by Escherichia coli upon expression of the prtM gene under the direction of the T7 RNA polymerase promoter was purified and used to obtain PrtM-specific antibodies. With these antibodies, immunogold labeling of lactococcal cells revealed that PrtM was associated with the lactococcal cell envelope. Western blot (immunoblot) analysis of whole lactococcal cells and isolated membrane vesicles indicated that PrtM was a membrane-associated protein. Radiolabeling of Lactococcus lactis with [3H]palmitic acid showed that PrtM was a lipoprotein. Partial secretion of PrtM into the culture medium was observed after Cys-24, the target residue for lipid modification, was replaced by an Ala residue by means of site-directed mutagenesis. This mutation did not affect proteinase activity.     

Safety and Immunogenicity of Neonatal Pneumococcal Conjugate Vaccination in Papua New Guinean Children: A Randomised Controlled Trial

Background

Approximately 826,000 children, mostly young infants, die annually from invasive pneumococcal disease. A 6-10-14-week schedule of pneumococcal conjugate vaccine (PCV) is efficacious but neonatal PCV may provide earlier protection and better coverage. We conducted an open randomized controlled trial in Papua New Guinea to compare safety, immunogenicity and priming for memory of 7-valent PCV (PCV7) given in a 0-1-2-month (neonatal) schedule with that of the routine 1-2-3-month (infant) schedule.

Methods

We randomized 318 infants at birth to receive PCV7 in the neonatal or infant schedule or no PCV7. All infants received 23-valent pneumococcal polysaccharide vaccine (PPV) at age 9 months. Serotype-specific serum IgG for PCV7 (VT) serotypes and non-VT serotypes 2, 5 and 7F were measured at birth and 2, 3, 4, 9, 10 and 18 months of age. Primary outcomes were geometric mean concentrations (GMCs) and proportions with concentration ≥0.35 µg/ml of VT serotype-specific pneumococcal IgG at age 2 months and one month post-PPV.

Results

We enrolled 101, 105 and 106 infants, respectively, into neonatal, infant and control groups. Despite high background levels of maternally derived antibody, both PCV7 groups had higher GMCs than controls at age 2 months for serotypes 4 (p<0.001) and 9V (p<0.05) and at age 3 months for all VTs except 6B. GMCs for serotypes 4, 9V, 18C and 19F were significantly higher (p<0.001) at age 2 months in the neonatal (one month post-dose2 PCV7) than in the infant group (one month post-dose1 PCV7). PPV induced significantly higher VT antibody responses in PCV7-primed than unprimed infants, with neonatal and infant groups equivalent. High VT and non-VT antibody concentrations generally persisted to age 18 months.

Conclusions

PCV7 is well-tolerated and immunogenic in PNG neonates and young infants and induces immunologic memory to PPV booster at age 9 months with antibody levels maintained to age 18 months.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00219401","term_id":"NCT00219401"}}NCT00219401{"type":"clinical-trial","attrs":{"text":"NCT00219401","term_id":"NCT00219401"}}NCT00219401     

Indirect immunofluorescence reaction of systemic and local humoral immunity in patients with inflammatory lung diseases. III. Secretory IgA antibodies to S. pneumoniae and H. influenzae autostrains

In acute pneumococcal infection with the clinical picture of acute bronchitis or the exacerbation of chronic bronchitis 88-93% of patients showed the formation of secretory IgA to S. pneumoniae autostrains, i.e. a high correlation between the occurrence of these bacteria in the sputum and the local immune response of the body. The characteristic feature of antipneumococcal secretory IgA is their earlier, more intensive and less prolonged formation in comparison with that of serum antibodies. During the exacerbation of chronic bronchitis only in 28% of the patients secretory IgA to H. influenzae autostrains and low correlation between the formation of secretory and serum antibodies were detected.     

Evaluation of test immunisation in the assessment of antibody deficiency syndromes.

Antibody responses after immunisation with pneumococcal polysaccharide did not correlate with the severity and frequency of infections in 22 patients with severe hypogammaglobulinaemia, when these were measured by a Farr radioimmunoassay. Five "healthy" patients with severe hypogammaglobulinaemia not only failed to make antipneumococcal polysaccharide antibody, when measured by radioimmunoassay, but also had very low or unrecordable antibody responses to Escherichia coli and failed to produce antibody when immunised with tetanus toxoid. Some of these subjects, however, did make small amounts of IgM antipneumococcal polysaccharide antibody when this was measured by an enzyme linked immunosorbent assay, while others retained some ability to produce IgM or IgA or both in their saliva. These findings show that the measurement of serum antibody responses after immunisation, with the possible exception of IgM antibodies to polysaccharides, is unlikely to be helpful in assessing the requirement for gammaglobulin replacement therapy in patients with hypogammaglobulinaemia.     

Localization and age-related changes in cytochrome P450 expression in APA hamster livers.

To establish the baseline data, age-related changes and the regional expression of the hepatic P450 isozymes in Syrian hamsters of the APA strain at 3, 6, 12, 18 months old were examined by immunological techniques. Immunohistochemical analysis of liver serial sections revealed that the midzonal and perivenous regions (zones 2 and 3, respectively) were stained with the anti-rat CYP1A1/2, 2B1/2 and 2E1 antibodies. These three antibodies most intensely stained the hepatocytes around the central vein. An anti-rat CYP3A2 staining section had a staining pattern with equally intense reactions in zones 2 and 3. On the other hand, CYP2C6, 2C11 and 4A1 were distributed diffusely throughout the hepatic acinus. There was no age-related difference in the expression pattern of any of the P450 isozymes examined. Total P450 content had a peak at 6 months of age and decreased to 60% of that level thereafter. Western-blot analysis revealed that the peak expressions of the isozymes detected with anti-rat CYP1A1/2, 2C6, 2E1 and 3A2 antibodies were observed in 6-month-old hamsters and declined in older ones. The CYP2B and 2C11 content reached the maximum at the age of 6 months and maintained almost the same level thereafter. The CYP4A level did not change from 3 to 6 months, and then declined to about 40% of the younger level at 12 and 18 months of age. These results suggest that the hepatic P450 isozymes of APA hamsters have region-specific expressions and most isozymes have their peaks of expression at 6 months of age, which differs from the patterns for rat P450.     

Active papain renatured and processed from insoluble recombinant propapain expressed in Escherichia coli.

For the first time the pro-form of a recombinant cysteine proteinase has been expressed at a high level in Escherichia coli. This inactive precursor can subsequently be processed to yield active enzyme. Sufficient protein can be produced using this system for X-ray crystallographic structure studies of engineered proteinases. A cDNA clone encoding propapain, a precursor of the papaya proteinase, papain, was expressed in E. coli using a T7 polymerase expression system. Insoluble recombinant protein was solubilized in 6 M guanidine hydrochloride and 10 mM dithiothreitol, at pH 8.6. A protein-glutathione mixed disulphide was formed by dilution into oxidized glutathione and 6 M GuHCl, also at pH 8.6. Final refolding and disulphide bond formation was induced by dilution into 3 mM cysteine at pH 8.6. Renatured propapain was processed to active papain at pH 4.0 in the presence of excess cysteine. Final processing could be inhibited by the specific cysteine proteinase inhibitors E64 and leupeptin, but not by pepstatin, PMSF or EDTA. This indicates that final processing was due to a cysteine proteinase and suggests that an autocatalytic event is required for papain maturation.     

Pneumococcal virulence factors: structure and function.

The overall goal for this review is to summarize the current body of knowledge about the structure and function of major known antigens of Streptococcus pneumoniae, a major gram-positive bacterial pathogen of humans. This information is then related to the role of these proteins in pneumococcal pathogenesis and in the development of new vaccines and/or other antimicrobial agents. S. pneumoniae is the most common cause of fatal community-acquired pneumonia in the elderly and is also one of the most common causes of middle ear infections and meningitis in children. The present vaccine for the pneumococcus consists of a mixture of 23 different capsular polysaccharides. While this vaccine is very effective in young adults, who are normally at low risk of serious disease, it is only about 60% effective in the elderly. In children younger than 2 years the vaccine is ineffective and is not recommended due to the inability of this age group to mount an antibody response to the pneumococcal polysaccharides. Antimicrobial drugs such as penicillin have diminished the risk from pneumococcal disease. Several pneumococcal proteins including pneumococcal surface proteins A and C, hyaluronate lyase, pneumolysin, autolysin, pneumococcal surface antigen A, choline binding protein A, and two neuraminidase enzymes are being investigated as potential vaccine or drug targets. Essentially all of these antigens have been or are being investigated on a structural level in addition to being characterized biochemically. Recently, three-dimensional structures for hyaluronate lyase and pneumococcal surface antigen A became available from X-ray crystallography determinations. Also, modeling studies based on biophysical measurements provided more information about the structures of pneumolysin and pneumococcal surface protein A. Structural and biochemical studies of these pneumococcal virulence factors have facilitated the development of novel antibiotics or protein antigen-based vaccines as an alternative to polysaccharide-based vaccines for the treatment of pneumococcal disease.     

Pneumococcal antibody concentrations and carriage of pneumococci more than 3 years after infant immunization with a pneumococcal conjugate vaccine

Background

A 9-valent pneumococcal conjugate vaccine (PCV-9), given in a 3-dose schedule, protected Gambian children against pneumococcal disease and reduced nasopharyngeal carriage of pneumococci of vaccine serotypes. We have studied the effect of a booster or delayed primary dose of 7-valent conjugate vaccine (PCV-7) on antibody and nasopharyngeal carriage of pneumococci 3–4 years after primary vaccination.

Methodology/Principal Findings

We recruited a subsample of children who had received 3 doses of either PCV-9 or placebo (controls) into this follow-up study. Pre- and post- PCV-7 pneumococcal antibody concentrations to the 9 serotypes in PCV-9 and nasopharyngeal carriage of pneumococci were determined before and at intervals up to 18 months post-PCV-7. We enrolled 282 children at a median age of 45 months (range, 38–52 months); 138 had received 3 doses of PCV-9 in infancy and 144 were controls. Before receiving PCV-7, a high proportion of children had antibody concentrations >0.35 µg/mL to most of the serotypes in PCV-9 (average of 75% in the PCV-9 and 66% in the control group respectively). The geometric mean antibody concentrations in the vaccinated group were significantly higher compared to controls for serotypes 6B, 14, and 23F. Antibody concentrations were significantly increased to serotypes in the PCV-7 vaccine both 6–8 weeks and 16–18 months after PCV-7. Antibodies to serotypes 6B, 9V and 23F were higher in the PCV-9 group than in the control group 6–8 weeks after PCV-7, but only the 6B difference was sustained at 16–18 months. There was no significant difference in nasopharyngeal carriage between the two groups.

Conclusions/Significance

Pneumococcal antibody concentrations in Gambian children were high 34–48 months after a 3-dose primary infant vaccination series of PCV-9 for serotypes other than serotypes 1 and 18C, and were significantly higher than in control children for 3 of the 9 serotypes. Antibody concentrations increased after PCV-7 and remained raised for at least 18 months.     

PiuA and PiaA, iron uptake lipoproteins of Streptococcus pneumoniae, elicit serotype independent antibody responses following human pneumococcal septicaemia

Streptococcus pneumoniae causes considerable morbidity and mortality worldwide. The need for a cheap and effective pneumococcal vaccine has necessitated the evaluation of common virulence-associated proteins as potential vaccine antigens. PiuA and PiaA are the lipoprotein components of two pneumococcal iron ABC transporters. Here, we show that patients with culture confirmed pneumococcal septicaemia have elevated levels of antibody to PiuA and PiaA in convalescent-phase, compared with acute-phase serum. Additionally, sera from septicaemic patients infected with 13 pneumococcal strains covering eight different serotypes, cross-reacted with recombinant PiuA-His(6) and PiaA-His(6) from a single pneumococcal strain, indicating that this immune response is serotype independent. Anti-PiuA and anti-PiaA antibodies were also found in healthy seven-month-old infants, indicating that they are immunogenic at a very early age.     

Role of spleen in immune response to polyvalent pneumococcal vaccine.

The immune response of lymphocytes to subcutaneously administered pneumococcal vaccine was studied in five patients without spleens and in five healthy subjects. Seven days after immunisation circulating B cells synthesising IgG antipneumococcal capsular polysaccharides (anti-PCP) appeared in both groups. Twenty one days after vaccination this B cell population had disappeared and a B cell subset which secreted IgM and IgG anti-PCP in the presence of pokeweed mitogen was detected in the normal but not in the splenectomised subjects. In the splenectomised group polyclonal IgM synthesis induced by pokeweed mitogen was defective. It was concluded that the early events of the immune response to PCP may be mediated by lymph nodes but that, later, the spleen acquires a central role in producing lymphocyte subsets capable of synthesising specific antibodies and that this might explain the increased sensitivity of splenectomised subjects to pneumococcal infection.     

Pneumococcal Virulence Factors: Structure and Function

The overall goal for this review is to summarize the current body of knowledge about the structure and function of major known antigens of Streptococcus pneumoniae, a major gram-positive bacterial pathogen of humans. This information is then related to the role of these proteins in pneumococcal pathogenesis and in the development of new vaccines and/or other antimicrobial agents. S. pneumoniae is the most common cause of fatal community-acquired pneumonia in the elderly and is also one of the most common causes of middle ear infections and meningitis in children. The present vaccine for the pneumococcus consists of a mixture of 23 different capsular polysaccharides. While this vaccine is very effective in young adults, who are normally at low risk of serious disease, it is only about 60% effective in the elderly. In children younger than 2 years the vaccine is ineffective and is not recommended due to the inability of this age group to mount an antibody response to the pneumococcal polysaccharides. Antimicrobial drugs such as penicillin have diminished the risk from pneumococcal disease. Several pneumococcal proteins including pneumococcal surface proteins A and C, hyaluronate lyase, pneumolysin, autolysin, pneumococcal surface antigen A, choline binding protein A, and two neuraminidase enzymes are being investigated as potential vaccine or drug targets. Essentially all of these antigens have been or are being investigated on a structural level in addition to being characterized biochemically. Recently, three-dimensional structures for hyaluronate lyase and pneumococcal surface antigen A became available from X-ray crystallography determinations. Also, modeling studies based on biophysical measurements provided more information about the structures of pneumolysin and pneumococcal surface protein A. Structural and biochemical studies of these pneumococcal virulence factors have facilitated the development of novel antibiotics or protein antigen-based vaccines as an alternative to polysaccharide-based vaccines for the treatment of pneumococcal disease.