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Original Article |

Virulence of Pneumococcal Proteins on the Inner Ear FREE

Patricia A. Schachern, BS; Vladimir Tsuprun, PhD; Sebahattin Cureoglu, MD; Patricia Ferrieri, MD; David E. Briles, PhD; Michael M. Paparella, MD; Steven Juhn, MD
[+] Author Affiliations

Author Affiliations: Departments of Otolaryngology (Ms Schachern and Drs Tsuprun, Cureoglu, Paparella, and Juhn) and Pediatrics and Laboratory Medicine and Pathology (Dr Ferrieri), University of Minnesota, Minneapolis; and Department of Microbiology, University of Alabama at Birmingham (Dr Briles).


Arch Otolaryngol Head Neck Surg. 2009;135(7):657-661. doi:10.1001/archotol.125.12.1371.
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Published online

Objective  To investigate the effects of the virulence characteristics of specific pneumococcal proteins on the inner ear.

Main Outcome Measures  A histologic comparison of inflammatory cell infiltration and pathologic changes in the round window membrane and inner ear.

Results  Most of the animals inoculated with high-dose pneumolysin or wild-type bacteria showed severe pathologic changes of the inner ears. The inner ears of most animals inoculated with surface protein A or surface antigen A–deficient bacteria appeared normal.

Conclusions  Pneumococcal surface protein A and pneumococcal surface antigen A are 2 important virulence factors in inner ear damage secondary to pneumococcal otitis media. Mutation of these virulence factors results in less inner ear damage.

Figures in this Article

Otitis media (OM) is one of the most common childhood diseases. Teele et al1 reported that 80% of children have 1 episode of OM by 3 years of age. In a study of antibiotic use and infections in infants and young children,2 OM was the most common indication and was responsible for 67.3% of antibiotics prescribed. The emergence of antibiotic-resistant bacterial strains has become a major health concern. Streptococcus pneumoniae is one of the important pathogens in acute OM, and its local rates of resistance have been reported to be from 50% to 60%.3

Antibiotic-resistant microorganisms have led to an increase in suppurative complications in OM.4 Redaelli de Zinis et al5 found middle-ear disease the most predisposing factor to sensorineural hearing loss and duration of disease to be related to its severity. Bacterial resistance to antibiotics has led to the development of alternatives such as the polyvalent vaccine. Although the Pneumovax 23 valent polysaccharide vaccine (Merck & Co, Whitehouse Station, New Jersey) is immunogenic and protective in most adults and children older than 5 years, it is not effective in children younger than 2 years,6 and conjugate vaccines have only limited serotype protection. This led to consideration of several pneumococcal proteins as alternative vaccine candidates; among these are pneumococcal surface protein A (PspA), pneumococcal surface antigen A (PsaA), and pneumolysin (Ply).

In a previous study, we demonstrated that pneumococci deficient in PspA (PspA) and PsaA proteins were less likely to pass from the middle ear through the round window membrane and into the inner ear than the wild-type or Ply-deficient (Ply) mutant bacterial strain (hereinafter, mutant).7 In this study, we compare inner-ear damage following high-dose and low-dose inoculations of wild-type mutant bacterial strains of these proteins into the middle ear.

Sixty-three young chinchillas weighing 250 to 350 g were included in this study. The care and use of these animals were approved by the Institutional Animal Care and Use Committee of the University of Minnesota, Minneapolis. All animals were anesthetized prior to bacterial inoculation with a combination of ketamine hydrochloride (100 mg/kg) and acepromazine maleate (10 mg/kg). They were inoculated bilaterally, with 0.5 mL of a high dose (105 to 107 colony-forming units [CFUs]) or low dose (103 CFUs) of S pneumoniae serotype 2 strain 39 (NCTC 7466) or its isogenic mutant deficient in PspA, PsaA mutant, that was derived by insertional inactivation mutagenesis of the PspA gene that was an insertional duplication derivate of the 7455 parent strain or the Ply mutant that had an insertionally inactivated Ply gene. Bacteria were grown in Todd-Hewitt broth (BD Diagnostics, Sparks, Maryland) containing yeast extract and plated on sheep blood agar. Erythromycin was added to plates for growth of the mutants, and bacteria were stored in glycerin freezing solution at −80°C. Bacteria were grown until they were in log phase, and their optical densities were measured. Concentrations were estimated, based on the specific strain, and diluted to the desired concentration. Ten-fold dilutions were plated, and viable cells were counted to confirm the actual concentration.

Two days after inoculation, animals were killed by overdose of sodium pentobarbital. The bullae were removed, and the middle-ear effusions grossly classified as clear or cloudy, yellow or white, and thick or thin. Cochleas were then gently perfused via the apex and oval window with glutaraldehyde, 2%, in 0.2M of phosphate buffer (pH, 7.4) for 2 hours. Samples were decalcified in ethylenediaminetetraacetic acid, 10%, for 3 days, washed in phosphate buffer, postfixed in osmium tetroxide, 1%, in 0.2M of phosphate buffer for 1 hour, and again washed with buffer. Following fixation, samples were dehydrated in a graded series of ethanol, followed by propylene oxide, and embedded in epoxy resin. Samples were cut at a thickness of 1 μm and stained with toluidine blue for light microscopic assessment. Samples for electron microscopy were cut at a thickness of 20 nm and stained with uranyl acetate and lead citrate.

Actual concentrations of bacteria for the animals inoculated with a high dose ranged from 7.5 × 105 to 3.4 × 107 CFUs for the wild-type strain, 2.3 × 106 to 5.8 × 107 CFUs for Ply, 7.5 × 105 to 8.1 × 107 CFUs for PspA mutants, and 5.5 × 105 to 5.5 × 106 CFUs for PsaA mutants. The condition of the animals is listed in Table 1. Of the 15 animals inoculated with the high-dose wild-type strain, 3 died, as did 1 of the animals inoculated with the high-dose PspA mutants. Three of the animals inoculated with the high-dose wild-type strain, and 1 of those inoculated with the PspA mutant, became extremely ill and had to be killed 1 day after inoculation. Actual concentrations of bacteria for the low-dose S pneumoniae were 4.8 × 103 to 7.5 × 103 CFUs for the wild-type strain and 2.8 × 103 CFUs for Ply, 0.6 × 103 CFUs for PspA, and 7.5 × 103 for PsaA strains. One of the chinchillas inoculated with low-dose PsaA died before the end of the experiment.

Table Graphic Jump LocationTable 1. Gross Characterization of Animals Following Inoculation With Wild-Type and Mutant Strains of Streptococcus pneumoniaea

All animals, those inoculated with both high-dose and low-dose bacterial strains, had middle-ear effusions that were generally thin. Thick purulent effusions were seen overlying the round window membranes in all 12 animals inoculated with the high-dose wild-type strain, in 4 of 9 inoculated with the Ply mutants, 5 of 9 inoculated with the PspA mutants, and in none of the 7 inoculated with the PsaA mutants. Thick purulent effusions were seen overlying the round window membranes in 4 of the 5 animal inoculated with the low-dose wild-type bacterial strain, 4 of 6 inoculated with the Ply mutants, 1 of 6 inoculated with the PspA mutants, and in none of the 3 inoculated with the PsaA mutants.

A summary of inner ear pathologic findings are listed in Table 2. Bacterial infiltration of the round window membranes (Figure 1) was seen in 6 of 12 animals inoculated with the high-dose wild-type strains, 7 of 9 animals inoculated with Ply mutants, 0 of 9 inoculated with PspA mutants, and 0 of 7 inoculated with PsaA mutants. Inflammatory cell infiltration of the round window membrane (Figure 1) was seen in 5 of 12 animals inoculated with the wild-type strain, 7 of 9 inoculated with Ply mutants, 2 of 9 inoculated with PspA mutants, and 1 of 7 inoculated with PsaA mutants. Bacterial infiltration of the scala tympani (Figure 2) was seen in 5 of 12 inoculated with wild-type strains, 7 of 9 inoculated with Ply mutants, in none inoculated with 9 PspA mutants, and 0 of 7 inoculated with PsaA mutants. Inflammatory cell infiltration of the scala tympani (Figure 2) was seen in 5 of 12 animals inoculated with the wild-type strain, 7 of 9 inoculated with Ply mutants, 3 of 9 inoculated with PspA mutants, and 1 of 7 inoculated with PsaA mutants. Strial edema (Figure 2) and/or strial atrophy occurred in 5 of 12 animals inoculated with the wild-type strain, 5 of 9 inoculated with Ply mutants, 2 of 9 inoculated with PspA mutants, and 1 of 7 inoculated with PsaA mutants. Damage to (Figure 2 and Figure 3) and/or loss of inner and outer hair cells occurred in 3 of 12 animals inoculated with the wild-type strain, 5 of 9 inoculated with Ply mutants, 2 of 9 inoculated with PspA mutants, and 1 of 7 inoculated with PsaA mutants. Pathologic changes to the neurons, including bacterial infiltration (Figure 3), hemorrhage, inflammatory cell infiltration, or neuronal damage, was seen in 4 of 12 animals inoculated with the wild-type strain, 6 of 9 inoculated with Ply mutants, 2 of 9 inoculated with PspA mutants, and 1 of 7 inoculated with PsaA mutants.

Place holder to copy figure label and caption
Figure 1.

Bacteria (arrows) can be seen in the round window membrane and scala tympani of this animal that was inoculated with high-dose pneumolysin-deficient mutant. ME indicates middle ear. Stained with toluidine blue; original magnification × 600.

Graphic Jump Location
Place holder to copy figure label and caption
Figure 2.

This animal inoculated with high-dose wild-type bacteria has edema of the stria vascularis (arrowhead) and hair cell damage (arrow). Stained with toluidine blue; original magnification × 600.

Graphic Jump Location
Place holder to copy figure label and caption
Figure 3.

Bacteria (arrows) and inflammatory cell infiltration can be seen in the scala tympani and within the osseous spiral lamina destroying the mylinated nerves. Note damage of hair cells. Stained with toluidine blue; original magnification × 600.

Graphic Jump Location
Table Graphic Jump LocationTable 2. Inner Ear Histopathologic Characteristicsa

In the animals inoculated with the low-dose bacterial strain, pathologic changes occurred only in those inoculated with the wild-type strain. Of the 5 animals inoculated with the wild-type strain, bacterial infiltration of the round window membrane and scala tympani was seen in 2, inflammatory cell infiltration of the round window membrane in 3, inflammatory cell infiltration of the scala tympani in 2, strial edema and/or atrophy in 1, hair cell damage and/or loss in 2, and neuronal damage in 1.

We have previously shown8 that bacteria and their toxins can pass from the middle ear into the inner ear in animals, resulting in inner ear damage and hearing loss. Hunter et al9 showed an association between high-frequency hearing loss and OM in children, even after middle-ear disease resolved and middle-ear dysfunction was excluded. A study5 of the consequences of chronic OM on inner ear function showed a correlation of the severity of sensorineural hearing loss with longer duration of middle-ear disease. A human temporal bone study of unilateral chronic OM found inner ear damage to be more severe in the ear with OM than in the contralateral ear.10

Widespread use of oral antibiotics has resulted in an alarming increase of antibiotic-resistant bacterial strains, increasing the potential for labyrinthitis, tympanogenic meningitis, and sensorineural hearing loss. Zapalac et al4 described 90 patients with suppurative complications of acute OM over a period of 7.5 years. They found that all but 2 cases of pneumococcal-related suppurative complications were attributable to resistant strains in the final 2.5 years, with resistant organisms isolated in 75% of patients.

Currently approved vaccines have proven to be only partially effective. The Pneumovax 23 valent polysaccharide vaccine does not protect children younger than 2 years,6 and conjugate vaccines have limited serotype protection.11 This led investigators to consider other potential vaccine candidates, such as pneumococcal proteins.

Pneumococcal proteins facilitate important aspects of pneumococcal colonization and/or invasion and can therefore serve as targets for the development of novel therapies to treat pneumococcal diseases. There are several pneumococcal proteins that contribute to the virulence and pathogenicity of streptococcus pneumoniae, including Ply, a cytoplasmic cholesterol binding, pore-forming protein12; PspA, a membrane-bound protein that prevents compliment fixation13,14; and PsaA, a surface protein that is a magnesium2+ and zinc2+ transporter, involved in growth and virulence.15

Although Ply is a potent cytotoxic agent, we found inner ear permeability and damage in animals inoculated with the Ply strains to be remarkably similar in virulence to the wild-type strain. Pneumolysin is located in the cytoplasm and is dependent on autolysin for its release. Sato et al16 showed that in the middle ear, pneumococcal OM pathogenesis is triggered principally by the inflammatory effects of intact and lytic cell wall products with, at most, a modest additional Ply effect.

Findings in the PspA and PsaA mutants were similar. Neither mutant seemed to pass through the round window membrane, and both mutants resulted in far less inner ear damage than either the wild-type strain or the Ply mutant. It must be acknowledged, however, that the animals were allowed to survive for only 2 days because those inoculated with the wild-type bacteria became sick and began to exhibit signs of labyrinthitis. It is possible that it may take the mutant bacteria longer to multiply to a level necessary to avoid clearance and enter the inner ear; however, the lack of virulence in these mutants may also be due to their missing proteins. The PspA protein is a cell surface protein that is protective against the host complement system.14,15 It also has the ability to bind lactoferrin,17 an iron storage glycoprotein predominantly found in mucosal secretions, including those of the middle ear,18 and a study19 has shown that resistance to pneumococcal carriage is dependent on mucosal rather than systemic immunity. The PspA mutants have also been shown to have a decreased capacity to form biofilms.20

Even though PspAs are antigenetically and structurally variable, they are immunologically cross-reactive,21,22 and immunization with a single PspA stimulates broadly cross-reactive antibodies.23 Because PspA is expressed by all clinically important capsular serotypes, it is particularly attractive as a vaccine candidate.24

Although clinically significant mucosal responses are directed to PspA, the systemic immune system seems to mount the highest humoral and cellular immune responses to PsaA.25 The PsaA protein is known to be a surface binding protein that inhibits complement activation.13 It has also been shown to play an important role in protection against pneumococcal carriage.26 In nasopharyngeal epithelial cells, adherence of S pneumoniae is the primary step essential for its pathogenesis,27 and Romero-Steiner et al28 identified a PsaA putative binding domain contained within a 28 amino acid peptide (P4) that contributes to PsaA adherence to nasopharyngeal cells. Immunization with a combination of PspA and PsaA proteins have been shown to offer better protection against nasal carriage in mice than immunization with either protein alone.27

Given the different roles of these proteins and the fact that they both seem to exhibit virulence factors that can have an impact on the inner ear, it may be interesting to explore a combination of these proteins in their role in the pathogenesis of OM and inner ear damage.

Correspondence: Patricia A. Schachern, BS, Room 226, Lions Research Building, 2001 Sixth St SE, Minneapolis, MN 55455 (schac002@tc.umn.edu).

Submitted for Publication: July 29, 2008; final revision received October 16, 2008; accepted November 13, 2008.

Author Contributions: All of the authors had full access to all the data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis. Study concept and design: Schachern, Tsuprun, Briles, Paparella, and Juhn. Acquisition of data: Schachern, Tsuprun, Cureoglu, and Ferrieri. Analysis and interpretation of data: Schachern and Tsuprun. Drafting of the manuscript: Schachern, Tsuprun, and Briles. Critical revision of the manuscript for important intellectual content: Schachern, Tsuprun, Cureoglu, Ferrieri, Briles, Paparella, and Juhn. Administrative, technical, and material support: Schachern, Tsuprun, Cureoglu, Ferrieri, and Briles. Study supervision: Ferrieri, Paparella, and Juhn.

Financial Disclosure: None reported.

Funding/Support: This study was supported by National Institute on Deafness and Other Communications Disorders (grant R01 DC006452), the International Hearing Foundation, the Starkey Foundation, and the Hubbard Foundation.

Teele  DWKlein  JORosner  B Epidemiology of otitis media during the first seven years of life in children in greater Boston: a prospective, cohort study. J Infect Dis 1989;160 (1) 83- 94
PubMed
Bergus  GRLevy  SMKirchner  HLWarren  JJLevy  BT A prospective study of antibiotic use and associated infections in young children. Paediatr Perinat Epidemiol 2001;15 (1) 61- 67
PubMed
McClay  JE Resistant bacteria in the adenoids: a preliminary report. Arch Otolaryngol Head Neck Surg 2000;126 (5) 625- 629
PubMed
Zapalac  JSBillings  KRSchwade  NDRoland  PS Suppurative complications of acute otitis media in the era of antibiotic resistance. Arch Otolaryngol Head Neck Surg 2002;128 (6) 660- 663
PubMed
Redaelli de Zinis  LOCampovecchi  CParrinello  GAntonelli  AR Predisposing factors for inner ear hearing loss association with chronic otitis media. Int J Audiol 2005;44 (10) 593- 598
PubMed
Paton  JC Novel pneumococcal surface proteins: role in virulence and vaccine potential. Trends Microbiol 1998;6 (3) 85- 87
PubMed
Schachern  PTsuprun  VCureoglu  C  et al.  The round window membrane in otitis media: effect of pneumococcal proteins. Arch Otolaryngol Head Neck Surg 2008;134 (6) 658- 662
PubMed
Schachern  PAPaparella  MMHybertson  RSano  SDuvall  AJ  III Bacterial tympanogenic labyrinthitis, meningitis, and sensorineural damage. Arch Otolaryngol Head Neck Surg 1992;118 (1) 53- 57
PubMed
Hunter  LLMargolis  RHRykken  JRLe  CTDaly  KAGiebink  GS High frequency hearing loss associated with otitis media. Ear Hear 1996;17 (1) 1- 11
PubMed
Cureoglu  SSchachern  PAPaparella  MMLindgren  BR Cochlear changes in chronic otitis media. Laryngoscope 2004;114 (4) 622- 626
PubMed
Hausdorff  WPBryant  JParadiso  PRSiber  GR Which pneumococcal serogroups cause the most invasive disease: implications for conjugate vaccine formulation and use, part I. Clin Infect Dis 2000;30 (1) 100- 121
PubMed
Roche  AMKing  SJWeiser  JN Live attenuated Streptococcus pneumoniae strains induce serotype-independent mucosal and systemic protection in mice. Infect Immun 2007;75 (5) 2469- 2475
PubMed
Tu  AHFulgham  RLMcCrory  MABriles  DESzalai  AJ Pneumococcal surface protein A inhibits complement activation by Streptococcus pneumoniae. Infect Immun 1999;67 (9) 4720- 4724
PubMed
Ren  BSzalai  AJHollingshead  SKBriles  DE Effects of PspA and antibodies to PspA on activation and deposition of complement on the pneumococcal surface. Infect Immun 2004;72 (1) 114- 122
PubMed
Dintilhac  AAlloing  GGranadel  CClaverys  JP Competence and virulence of Streptococcus pneumoniae: Adc and PsaA mutants exhibit a requirement for Zn and Mn resulting from inactivation of putative ABC metal permeases. Mol Microbiol 1997;25 (4) 727- 739
PubMed
Sato  KQuartey  MKLiebeler  CLLe  CTGiebink  GS Roles of autolysin and pneumolsin in middle ear inflammation caused by a type 3 streptococcus pneumoniae strain in the chinchilla otitis media model. Infect Immun 1996;64 (4) 1140- 1145
PubMed
Jedrzejas  MJ Unveiling molecular mechanisms of pneumococcal surface protein A interaction with antibodies and lactoferrin. Clin Chim Acta 2006;367 (1-2) 1- 10
PubMed
Giebink  GSCarlson  BAHetherington  SVHostetter  MKLe  CTJuhn  SK Bacterial and polymorphonuclear leukocyte contribution to middle ear inflammation in chronic otitis media with effusion. Ann Otol Rhinol Laryngol 1985;94 (4, pt 1) 398- 402
PubMed
Wu  HYNahm  MHGuo  YRussell  MWBriles  DE Intranasal immunization of mice with PspA (pneumococcal surface protein A) can prevent intranasal carriage, pulmonary infection, and sepsis with Streptococcus pneumoniae. J Infect Dis 1997;175 (4) 839- 846
PubMed
Moscoso  MGarcia  ELopez  R Biofilm formation by Streptococcus pneumoniae: role of choline, extracellular DNA, and capsular polysaccharide in microbial accretion. J Bacteriol 2006;188 (22) 7785- 7795
PubMed
Crain  MJWaltman  WD  IITurner  JS  et al.  Pneumococcal surface protein A (PspA) is serologically highly variable and is expressed by all clinically important capsular serotypes of Streptococcus pneumoniaeInfect Immun 1990;58 (10) 3293- 3299
PubMed
Palaniappan  RSingh  SSingh  UP  et al.  Differential PsaA-, PspA-, PspC-, and Pdb-specific immune responses in a mouse model of pneumococcal carriage. Infect Immun 2005;73 (2) 1006- 1013
PubMed
Nabors  GSBraun  PAHerrmann  DJ  et al.  Immunization of healthy adults with a single recombinant pneumococcal surface protein A (PspA) variant stimulates broadly cross-reactive antibodies to heterologous PspA molecules. Vaccine 2000;18 (17) 1743- 1754
PubMed
Briles  DEHollingshead  SBrooks-Walter  A  et al.  The potential to use PspA and other pneumococcal proteins to elicit protection against pneumococcal infection. Vaccine 2000;18 (16) 1701- 1711
PubMed
Arulanandam  BPLynch  JMBriles  DEHollingshead  SMetzger  DW Intranasal vaccination with pneumococcal surface protein A and interleukin-12 augments antibody-mediated opsonization and protective immunity against Streptococcus pneumoniae infection. Infect Immun 2001;69 (11) 6718- 6724
PubMed
Tuomanen  E Molecular and cellular biology of pneumococcal infection. Curr Opin Microbiol 1999;2 (1) 35- 39
PubMed
Briles  DEAdes  EPaton  JC  et al.  Intranasal immunization of mice with a mixture of the pneumococcal proteins PsaA and PspA is highly protective against nasopharyngeal carriage of Streptococcus pneumoniaeInfect Immun 2000;68 (2) 796- 800
PubMed
Romero-Steiner  SCaba  JRajam  G  et al.  Adherence of recombinant pneumococcal surface adhesin A (rPsaA)-coated particles to human nasopharyngeal epithelial cells for the evaluation of anti-PsaA functional antibodies. Vaccine 2006;24 (16) 3224- 3231
PubMed

Figures

Place holder to copy figure label and caption
Figure 1.

Bacteria (arrows) can be seen in the round window membrane and scala tympani of this animal that was inoculated with high-dose pneumolysin-deficient mutant. ME indicates middle ear. Stained with toluidine blue; original magnification × 600.

Graphic Jump Location
Place holder to copy figure label and caption
Figure 2.

This animal inoculated with high-dose wild-type bacteria has edema of the stria vascularis (arrowhead) and hair cell damage (arrow). Stained with toluidine blue; original magnification × 600.

Graphic Jump Location
Place holder to copy figure label and caption
Figure 3.

Bacteria (arrows) and inflammatory cell infiltration can be seen in the scala tympani and within the osseous spiral lamina destroying the mylinated nerves. Note damage of hair cells. Stained with toluidine blue; original magnification × 600.

Graphic Jump Location

Tables

Table Graphic Jump LocationTable 1. Gross Characterization of Animals Following Inoculation With Wild-Type and Mutant Strains of Streptococcus pneumoniaea
Table Graphic Jump LocationTable 2. Inner Ear Histopathologic Characteristicsa

References

Teele  DWKlein  JORosner  B Epidemiology of otitis media during the first seven years of life in children in greater Boston: a prospective, cohort study. J Infect Dis 1989;160 (1) 83- 94
PubMed
Bergus  GRLevy  SMKirchner  HLWarren  JJLevy  BT A prospective study of antibiotic use and associated infections in young children. Paediatr Perinat Epidemiol 2001;15 (1) 61- 67
PubMed
McClay  JE Resistant bacteria in the adenoids: a preliminary report. Arch Otolaryngol Head Neck Surg 2000;126 (5) 625- 629
PubMed
Zapalac  JSBillings  KRSchwade  NDRoland  PS Suppurative complications of acute otitis media in the era of antibiotic resistance. Arch Otolaryngol Head Neck Surg 2002;128 (6) 660- 663
PubMed
Redaelli de Zinis  LOCampovecchi  CParrinello  GAntonelli  AR Predisposing factors for inner ear hearing loss association with chronic otitis media. Int J Audiol 2005;44 (10) 593- 598
PubMed
Paton  JC Novel pneumococcal surface proteins: role in virulence and vaccine potential. Trends Microbiol 1998;6 (3) 85- 87
PubMed
Schachern  PTsuprun  VCureoglu  C  et al.  The round window membrane in otitis media: effect of pneumococcal proteins. Arch Otolaryngol Head Neck Surg 2008;134 (6) 658- 662
PubMed
Schachern  PAPaparella  MMHybertson  RSano  SDuvall  AJ  III Bacterial tympanogenic labyrinthitis, meningitis, and sensorineural damage. Arch Otolaryngol Head Neck Surg 1992;118 (1) 53- 57
PubMed
Hunter  LLMargolis  RHRykken  JRLe  CTDaly  KAGiebink  GS High frequency hearing loss associated with otitis media. Ear Hear 1996;17 (1) 1- 11
PubMed
Cureoglu  SSchachern  PAPaparella  MMLindgren  BR Cochlear changes in chronic otitis media. Laryngoscope 2004;114 (4) 622- 626
PubMed
Hausdorff  WPBryant  JParadiso  PRSiber  GR Which pneumococcal serogroups cause the most invasive disease: implications for conjugate vaccine formulation and use, part I. Clin Infect Dis 2000;30 (1) 100- 121
PubMed
Roche  AMKing  SJWeiser  JN Live attenuated Streptococcus pneumoniae strains induce serotype-independent mucosal and systemic protection in mice. Infect Immun 2007;75 (5) 2469- 2475
PubMed
Tu  AHFulgham  RLMcCrory  MABriles  DESzalai  AJ Pneumococcal surface protein A inhibits complement activation by Streptococcus pneumoniae. Infect Immun 1999;67 (9) 4720- 4724
PubMed
Ren  BSzalai  AJHollingshead  SKBriles  DE Effects of PspA and antibodies to PspA on activation and deposition of complement on the pneumococcal surface. Infect Immun 2004;72 (1) 114- 122
PubMed
Dintilhac  AAlloing  GGranadel  CClaverys  JP Competence and virulence of Streptococcus pneumoniae: Adc and PsaA mutants exhibit a requirement for Zn and Mn resulting from inactivation of putative ABC metal permeases. Mol Microbiol 1997;25 (4) 727- 739
PubMed
Sato  KQuartey  MKLiebeler  CLLe  CTGiebink  GS Roles of autolysin and pneumolsin in middle ear inflammation caused by a type 3 streptococcus pneumoniae strain in the chinchilla otitis media model. Infect Immun 1996;64 (4) 1140- 1145
PubMed
Jedrzejas  MJ Unveiling molecular mechanisms of pneumococcal surface protein A interaction with antibodies and lactoferrin. Clin Chim Acta 2006;367 (1-2) 1- 10
PubMed
Giebink  GSCarlson  BAHetherington  SVHostetter  MKLe  CTJuhn  SK Bacterial and polymorphonuclear leukocyte contribution to middle ear inflammation in chronic otitis media with effusion. Ann Otol Rhinol Laryngol 1985;94 (4, pt 1) 398- 402
PubMed
Wu  HYNahm  MHGuo  YRussell  MWBriles  DE Intranasal immunization of mice with PspA (pneumococcal surface protein A) can prevent intranasal carriage, pulmonary infection, and sepsis with Streptococcus pneumoniae. J Infect Dis 1997;175 (4) 839- 846
PubMed
Moscoso  MGarcia  ELopez  R Biofilm formation by Streptococcus pneumoniae: role of choline, extracellular DNA, and capsular polysaccharide in microbial accretion. J Bacteriol 2006;188 (22) 7785- 7795
PubMed
Crain  MJWaltman  WD  IITurner  JS  et al.  Pneumococcal surface protein A (PspA) is serologically highly variable and is expressed by all clinically important capsular serotypes of Streptococcus pneumoniaeInfect Immun 1990;58 (10) 3293- 3299
PubMed
Palaniappan  RSingh  SSingh  UP  et al.  Differential PsaA-, PspA-, PspC-, and Pdb-specific immune responses in a mouse model of pneumococcal carriage. Infect Immun 2005;73 (2) 1006- 1013
PubMed
Nabors  GSBraun  PAHerrmann  DJ  et al.  Immunization of healthy adults with a single recombinant pneumococcal surface protein A (PspA) variant stimulates broadly cross-reactive antibodies to heterologous PspA molecules. Vaccine 2000;18 (17) 1743- 1754
PubMed
Briles  DEHollingshead  SBrooks-Walter  A  et al.  The potential to use PspA and other pneumococcal proteins to elicit protection against pneumococcal infection. Vaccine 2000;18 (16) 1701- 1711
PubMed
Arulanandam  BPLynch  JMBriles  DEHollingshead  SMetzger  DW Intranasal vaccination with pneumococcal surface protein A and interleukin-12 augments antibody-mediated opsonization and protective immunity against Streptococcus pneumoniae infection. Infect Immun 2001;69 (11) 6718- 6724
PubMed
Tuomanen  E Molecular and cellular biology of pneumococcal infection. Curr Opin Microbiol 1999;2 (1) 35- 39
PubMed
Briles  DEAdes  EPaton  JC  et al.  Intranasal immunization of mice with a mixture of the pneumococcal proteins PsaA and PspA is highly protective against nasopharyngeal carriage of Streptococcus pneumoniaeInfect Immun 2000;68 (2) 796- 800
PubMed
Romero-Steiner  SCaba  JRajam  G  et al.  Adherence of recombinant pneumococcal surface adhesin A (rPsaA)-coated particles to human nasopharyngeal epithelial cells for the evaluation of anti-PsaA functional antibodies. Vaccine 2006;24 (16) 3224- 3231
PubMed

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The American Medical Association is accredited by the Accreditation Council for Continuing Medical Education to provide continuing medical education for physicians. The AMA designates this journal-based CME activity for a maximum of 1 AMA PRA Category 1 CreditTM per course. Physicians should claim only the credit commensurate with the extent of their participation in the activity. Physicians who complete the CME course and score at least 80% correct on the quiz are eligible for AMA PRA Category 1 CreditTM.
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For CME Course: A Proposed Model for Initial Assessment and Management of Acute Heart Failure Syndromes
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Citing articles are presented as examples only. In non-demo SCM6 implementation, integration with CrossRef’s "Cited By" API will populate this tab (http://www.crossref.org/citedby.html).
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