Pneumococcal acute otitis media in children

Objectives   To evaluate the clinical and laboratory findings of Streptococcus pneumoniae acute otitis media in children during a 1 year period.
Methods   From October 1995 to September 1996, 113 children aged 2 months to 14 years (median 18 months), with S. pneumoniae acute otitis media were studied. Susceptibility testing was performed by the Kirby-Bauer method and the E- test, and serotyping by the Quellung reaction.
Results   E- test assays detected five isolates (4.4%) to be highly resistant to penicillin and 13 (11.5%) that had intermediate resistance. All isolates were found to be susceptible to vancomycin, rifampicin and cefotaxime. In total, 25 isolates (22.1%) were resistant to one or more drugs. Fifty per cent of the penicillin-resistant or intermediately resistant S. pneumoniae isolates were resistant to multiple drugs, whereas only 2.1% of the penicillin-susceptible isolates were resistant to multiple drugs. The predominating serogroups of the isolates with reduced susceptibility to penicillin were the 19 (61.1%), 9 (16.7%), 23 (11.1%), 6 (5.5%) and 14 (5.5%) whereas those of the susceptible isolates were the 19 (26.3%), 14 (13.7%), 3 (11.6%), 6 (11.6%), 9 (8.4%), 1 (5.3%) and 12 (5.3%).
Conclusions   Streptococcus pneumoniae isolates from children with acute otitis media were penicillin-insensitive in 15.9%. The multiresistant S. pneumoniae isolates belonged to serogroups: 19 (45.4%), 9 (27.3%), 6 (18.2%) and 23 (9.1%).     

Nasopharyngeal secretory immunoglobulins in children with recurrent acute otitis media and secretory otitis media

Secretory IgA (SIgA) and secretory IgM (SIgM), total IgA and total IgM were measured in plasma and nasopharyngeal secretions (NPS) from young children with different degrees of otitis proneness. Significantly higher levels of plasma IgM and lower levels of NPS-SIgM were found in children with recurrent episodes of acute otitis media (rAOM) compared with children suffering from secretory otitis media (SOM) and healthy controls. Both plasma IgA and NPS-SIgA were evenly distributed in the three groups of children investigated, and in most children the levels of NPS-SIgA exceeded plasma IgA levels. Plasma SIgA was significantly increased in children with rAOM and SOM, probably resulting from frequent occurrence of inflammatory events at the nasopharyngeal level. No correlation could be demonstrated between NPS-SIgA and plasma IgA, or between NPS-SIgM and plasma IgM. Also, for both NPS-SIgA and NPS-SIgM, there was no correlation with age. A negative correlation was observed between the transudation index of albumin to the nasopharynx and the ratio of NPS-SIgA to total NPS-IgA. A ratio of 1 (100%) corresponded to a transudation index of 8%. The ratios of NPS-SIgA to total NPS-IgA varied considerably and a range of 39%-88% could only to some extent be explained by transudation of plasma IgA to NPS. The results of the present study show that the children with rAOM and SOM are well furnished with locally produced SIgA antibodies at the nasopharyngeal level. In children with SOM, the nasopharyngeal hypofunction in the case of low NPS-SIgM seems to be less pronounced compared with that of otitis-prone children.     

Antipolysaccharide antibodies in 450 children with otitis media

We have measured antibodies to pneumococcal and Haemophilus polysaccharides in a prospective study of 450 children aged 2–16 years with otitis media requiring grommets (ear tubes). Pneumococcal antibody levels were significantly higher in the 2–6 year (P < 0.004) and 7–10 year (P < 0.04) study groups in comparison with age-matched controls. There was no difference in Haemophilus antibody levels between the study and control group children for the age groups 2–6 years and 11–16 years. Haemophilus antibody levels were significantly lower in the 7–10 year (P < 0.003) group in comparison with age-matched controls. Eighty-eight out of 450 (19.6%) children had pneumococcal antibody levels below the 25th percentile. Nineteen out of 88 (21.6%) children with pneumococcal antibody levels below the 25th centile were test immunized with 23 valent Pneumococcal polysaccharide and unconjugated Haemophilus type b capsular polysaccharide. Of these 19 children (aged 4–11 years), five mounted suboptimal responses to both polysaccharide antigens, whilst one child failed to respond to Haemophilus polysaccharide alone. There was no significant difference in the prevalence of IgG subclass deficiency between the normal responders and poor responders to immunization (P= 0.12). We found no evidence of specific polysaccharide antibody deficiency in the vast majority of the 450 children studied. However, the significance of poor antibody responses to test immunization in a small minority of children with otitis media is unclear. Long-term follow up of these children is required to determine whether poor immunization responses herald the development of frank antibody deficiency.     

Plasma IgG, IgG subclasses and acute-phase proteins in children with recurrent acute otitis media

Quantitation of IgG, IgG subclasses and acute-phase proteins was performed in plasma samples from 156 children, aged 2 to 162 months, with varying degrees of otitis proneness. None of the children had acute otitis media or had received any antibiotics 3 weeks before the examination. Children with recurrent acute otitis media (rAOM) had significantly higher levels of total IgG, IgG1, and significantly lower levels of IgG2 than healthy children (p less than 0.02, p less than 0.0003, and p less than 0.03, respectively). However, the low levels of IgG2 found in the rAOM children could at least, to some extent, be explained by the observation that these children were somewhat younger than the healthy children. Except for recurrent episodes of common colds, children suffering from secretory otitis media (SOM) most often show no clinical signs of inflammatory events. Nevertheless, children with SOM had raised levels of plasma IgG1, indicating recurrent polyclonal stimulation of the immune apparatus, which seems to be less pronounced than that of rAOM children. Levels of acute-phase proteins, haptoglobin, orosomucoid and alpha 1-antitrypsin were evenly distributed in the children investigated, reflecting that they had no acute illness at the time of plasma sampling.     

Protection by serum antibodies in experimental nontypable Haemophilus influenzae otitis media.

The chinchilla experimental model of otitis media was used to examine the importance of serum antibodies in protection against disease caused by nontypable Haemophilus influenzae. An immune serum pool was prepared by immunizing chinchillas with killed bacterial cells of nontypable H. influenzae 3245. Pooled preimmune or immune serum from these immunized animals was administered intravenously to a group of nonimmune chinchillas 1 day before intrabullar challenge with strain 3245. Of 5 animals receiving preimmune serum, 5 developed otitis media compared with 0 of 10 animals receiving immune serum (P = 0.008). The immune serum pool contained antibodies directed against both surface-exposed outer membrane proteins and lipopolysaccharide (LPS). The 39-kilodalton major outer membrane protein was the immunodominant surface protein. Anti-LPS antibodies were removed from the immune serum pool by affinity chromatography, and affinity-purified anti-LPS antibodies were recovered. Immune serum, immune serum absorbed of LPS antibodies, or affinity-purified LPS antibodies were then administered to another group of experimental animals 1 day before bacterial challenge. Of four animals that received the affinity-purified LPS antibodies, four developed otitis compared with zero of four animals that received the immune serum or zero of four animals that received the LPS-absorbed immune serum (P = 0.028). These studies indicate that passive immunization with immune serum is protective in experimental nontypable H. influenzae otitis media and that bacterial outer membrane proteins may be the principal targets of protective antibody.     

Characterization of ear fluid isolates of Alloiococcus otitidis from patients with recurrent otitis media.

Nineteen isolates of Alloiococcus otitidis from ear fluid samples collected by tympanostomy from patients at four geographic locations were identified by phenotypic characterization and genetic relatedness. Initial growth of A. otitidis isolates occurred after 3 days at 37 degrees C on brain heart infusion (BHI) agar with 5% rabbit blood. Heavy growth occurred in BHI broth supplemented with 0.07% lecithin and 0.5% Tween 80 after 4 days of incubation. The isolates were gram-positive cocci that divided on an irregular plane and produced metabolic lactic acid, pyrrolidonyl arylamidase, and leucine aminopeptidase. These cocci grew sparsely in 6.5% NaCl-BHI broth, were asaccharolytic on both fermentative and oxidative bases, and were cytochrome negative by the iron-porphyrin test. The cellular fatty acid profile of A. otitidis was distinguished from those of related genera and characterized by major amounts ( > or = 14%) of 16:0, 18:2, 18:1 omega 9c, and 18:0 and smaller amounts of 14:0, 16:1 omega 7c, 17:0, and 18:1 omega 7c. Fifteen isolates demonstrated > 69% relatedness by DNA-DNA hybridization. Four isolates plus the original 15 were confirmed as A. otitidis by dot blot hybridization with a digoxigenin-labeled nucleotide probe specific for this species. The intergenic space between the genes coding for the 16S and 23S rRNAs of alloiococci was amplified by PCR, analyzed by restriction fragment length polymorphism, and determined to consist of three different genetic types. Although beta-lactamase negative, A. otitidis demonstrated intermediate levels of resistance to beta-lactams, including expanded-spectrum cephalosporins, and were resistant to trimethoprim-sulfamethoxazole and erythromycin.     

Recent advances in otitis media with effusion

OME is similar to other chronic inflammatory diseases with the additional fact that ventilation of the middle ear is an extremely important and necessary part of therapy. Antimicrobial therapy alone may not cure middle ear inflammatory disease as long as Eustachian tube dysfunction persists. If Eustachian tube dysfunction and middle ear fluid persists, all forms of immunologic reactivity may occur in the middle ear which may result in persistent effusion and damage to the mucosal system, to the middle ear ossicles, and to the mastoid mucosal system. These changes may require more aggressive surgical management. It is obvious that we must learn more about the physiology and pathophysiology of the Eustachian tube; about understanding of the systemic, and particularly the local humoral and cell-mediated immune response in the middle ear. Finally, we must determine the potential role of inflammatory mediators, both in the protection against disease and also in the potential role that they play in tissue injury. Encouraging progress is being made in all of these areas and it seems likely that the increased research interest and productivity from laboratories throughout the world will eventually lead to the goal of enhanced prevention and improved treatment of OME.     

Molecular epidemiology of pneumococcal colonization in response to pneumococcal conjugate vaccination in children with recurrent acute otitis media

A randomized double-blind trial with a 7-valent pneumococcal conjugate vaccine was conducted in The Netherlands among 383 children, aged 1 to 7 years, with a history of recurrent acute otitis media. No effect of vaccination on the pneumococcal colonization rate was found. However, a shift in serotype distribution was clearly observed (R. Veenhoven et al., Lancet 361:2189-2195, 2003). We investigated the molecular epidemiology of 921 pneumococcal isolates retrieved from both the pneumococcal vaccine (PV) and control vaccine (CV) groups during the vaccination study. Within individuals a high turnover rate of pneumococcal restriction fragment end labeling genotypes, which was unaffected by vaccination, was observed. Comparison of the genetic structures before and after completion of the vaccination scheme revealed that, despite a shift in serotypes, there was clustering of 70% of the pneumococcal populations. The remaining isolates (30%) were equally observed in the PV and CV groups. In addition, the degree of genetic clustering was unaffected by vaccination. However, within the population genetic structure, nonvaccine serotype clusters with the serotypes 11, 15, and 23B became predominant over vaccine-type clusters after vaccination. Finally, overall pneumococcal resistance was low (14%), and, albeit not significant, a reduction in pneumococcal resistance as a result of pneumococcal vaccination was observed. Molecular surveillance of colonization in Dutch children shows no effect of pneumococcal conjugate vaccination on the degree of genetic clustering and the genetic structure of the pneumococcal population. However, within the genetic pneumococcal population structure, a clear shift toward nonvaccine serotype clusters was observed.     

Cytolytic complement activity in otitis media with effusion

Otitis media with effusion (OME) is a chronic inflammation persisting in the middle ear cavity of at least 8 weeks duration. Middle ear effusion (MEE; n = 38), samples from children suffering from OME were investigated for their direct cytolytic activity or an ability to enhance complement lysis of unsensitized bystander cells. Thirteen of the 38 MEEs had direct endogenous haemolytic activity and 27 samples had an ability to enhance serum-initiated lysis. Using an enzyme immunoassay, high levels of terminal complement complexes (TCC) were detected in the MEE samples (mean 34.1 microg/ml, range 5--89 microg/ml). This indicated strong local complement activation that had progressed to the terminal stage. As one potential factor promoting complement activation we identified both monomeric and trimeric properdin in MEE by Western blotting. By stabilizing C3 and C5 convertases properdin accelerates the alternative and terminal pathways of complement. On the other hand, the membrane attack complex (MAC) inhibitor CD59, which was found to be extensively shed into the MEE in a functionally active form, may control excessive cytotoxicity of the MEE. In conclusion, intense complement activation, up to the terminal level, maintains ongoing inflammation in the middle ear cavity and can pose a threat to the local epithelium.     

Properdin deficiency associated with recurrent otitis media and pneumonia, and identification of male carrier with Klinefelter syndrome

Properdin is an initiator and stabilizer of the alternative complement activation pathway (AP). Deficiency of properdin is a rare X-linked condition characterized by increased susceptibility to infection with Neisseria meningitidis associated with a high mortality rate. We report properdin deficiency in a large Pakistani family. The index cases were found by screening for immunodeficiency due to a history of recurrent infections. This revealed absent AP activity, but normal classical and lectin pathway activity. Sequencing of the properdin gene (PFC) revealed a novel frameshift mutation. When all available relatives (n = 24) were screened for the mutation, four affected males, four female carriers and a male heterozygous carrier were identified. He was subsequently diagnosed with Klinefelter syndrome. A questionnaire revealed a striking association between properdin deficiency and recurrent otitis media (P = 0.0012), as well as recurrent pneumonia (P = 0.0017).This study is the first to show a significant association between properdin deficiency and recurrent infections.     

Local production of IgG to pneumococcal C-polysaccharide in upper airway secretions from children with recurrent acute otitis media

Antibodies against C-polysaccharide (C-Ps), a common cell wall component of all pneumococci, may be of importance for the elimination of decaying pneumococci. By means of ELISA with phenylated C-Ps, anti-C-Ps IgG was measured in samples of plasma and upper airway secretions from otitis-prone children (OP), children with fewer episodes of recurrent acute otitis media (rAOM), and children with no previous history of AOM, but suffering from secretory otitis media (SOM). All children were free from acute illness at the time of sampling. No statistically significant differences of anti-C-Ps IgG in plasma or in nasopharyngeal secretions (NPS) were found between any of the groups. Based on calculations of the correlation between levels of anti-C-Ps IgG in plasma and NPS, and of the transudation ratios of anti-C-Ps IgG, total IgG, and albumin from plasma to NPS, we suggest that a significant amount of the anti-C-Ps IgG in NPS must be locally produced. The additional finding that OP children had significantly higher levels of anti-C-Ps IgG in their middle ear effusions (MEE) than SOM children points in the same direction. Anti-C-Ps IgA and IgM were detected in very low concentrations in plasma and secretion samples.