Microdialysis studies of the middle ear distribution kinetics of amoxicillin in the awake chinchilla.

This work was performed to develop an experimental animal model for the study of antibiotic drug distribution into middle ear fluid (MEF) and to evaluate its relevance and significance to the clinical treatment of otitis media (OM). Chinchillas were assigned to normal or infected ear groups after Eustachian tube obstruction (ETO) or direct trans-bullar inoculation with type 3 Streptococcus pneumoniae. Following survival surgery to implant microdialysis (MD) probes in the jugular vein and middle ear (ME), amoxicillin was given intravenously (iv) as a bolus or infusion. Drug concentrations in blood and MEF were continuously monitored by microdialysis. The measured concentrations were corrected for probe recovery by simultaneous retrodialysis. Multiple MEF and blood sampling was also performed to validate the animal model and MD sampling technique. Bacterial infection was successfully induced 3-7 days after the inoculation, whereas the control group gave negative bacterial culture results. The beta-lactam antibiotic, amoxicillin, exhibited an elimination half-life of 0.33+/-0.23 h (n = 9) in chinchilla blood, 1.46+/-0.50 h (n = 5) and 1.75+/-0.84 h (n = 4) in MEF of normal and infected ears (p = 0.6), respectively. MEF-to-blood amoxicillin concentration ratios at steady state following iv infusion were 0.26+/-0.06 (n = 5) and 0.28+/-0.11 (n = 4) for normal and infected ears (p = 0.7), respectively. MD allows continuous monitoring of drug concentration-time profiles in blood and MEF in an awake chinchilla model. The concentrations measured by MD were validated by direct sampling. The ratio of the area under the curve (AUC) of drug concentration in MEF versus time to that in blood after iv bolus doses was less than unity, as was the steady-state concentration ratio following constant-rate iv infusion, suggesting an active transport mechanism was involved in the efflux of amoxicillin from the ME of chinchilla. The results of studies involving infected ears were not significantly different from those in normal ears in terms of amoxicillin distribution across the ME mucosal membrane after systemic administration.

A multi-laboratory evaluation of an enzyme-linked immunoassay quantitating human antibodies to Streptococcus pneumoniae polysaccharides.

An enzyme-linked immunoassay (EIA) is described and evaluated which quantitates human antibodies to serotype specific S. pneumoniae polysaccharide (PnPs) in human sera. Based on the observations previously described by Koskela (1), native PnPs are used as coating antigens and sera are absorbed with a soluble pneumococcal absorbant material containing C-polysaccharide (CPs) to ensure measurement of serotype specific anti-PnPs antibodies. The robustness of this method was evaluated by ten laboratories using the same reagents, protocol, and five human serum samples. Reproducible antibody values were obtained for IgM, IgG, and IgA antibodies to five different PnPs serotypes, 3, 6B, 14, 19F, and 23F. The overall mean percent coefficients of variation in this interlaboratory study for all five selotype specific anti-PnPs determinations with the five coded sera were 30% for IgG, 3/% for IgM, and 36% for IgA. This assay can be standardized for quantitation of serotype specific anti-PnPs antibodies, allowing comparison of antibody values in vaccine trials evaluating pneumococcal vaccines.

Low cord blood pneumococcal antibody concentrations predict more episodes of otitis media.

OBJECTIVE: To determine if cord blood anticapsular polysaccharide pneumococcal IgG antibody concentration was related to the number of otitis media (OM) and acute OM episodes during the first year of life. DESIGN: Prospective study following infants from birth to 24 months. SETTING: Health maintenance organization. PATIENTS: The study population consisted of 415 infants whose mothers volunteered for the study during pregnancy. Cord blood samples were collected and infants were followed up for OM in the health maintenance organization. Ninety-seven percent of the infants were white, 49% male, 3% from households with annual incomes of less than $20 000, and 30% from households with annual incomes of more than $60 000. MAIN OUTCOME MEASURE: Number of physician-diagnosed OM episodes, including both OM with effusion and acute OM, and acute OM episodes from birth to 12 months. RESULTS: With univariate analysis, low cord blood antibody concentrations against serotypes 3 and 19F predicted more acute OM episodes (P =.04 and P =.05, respectively), and low antibody concentrations against serotypes 19F and 23F predicted more OM episodes (P =.04 and P =.05, respectively) over the first year of life. With Poisson regression, which adjusted for variables related to the recurrence of OM and having low cord blood antibody concentrations, serotype 19F remained significantly related to the number of OM episodes (relative risk for lowest quartiles vs upper 3 quartiles 1.23; 95% confidence interval, 1.02-1.50; P =.03). CONCLUSIONS: Low cord blood antibody concentrations to serotype 19F predicted more OM episodes over the first 12 months of life. These results suggest the potential benefit of maternal immunization to raise neonatal antipolysaccharide pneumococcal antibody concentration and delay the onset and reduce the number of OM episodes.

Low cord blood type 14 pneumococcal IgG1 but not IgG2 antibody predicts early infant otitis media.

Type-specific IgG1 and IgG2 antibodies to Streptococcus pneumoniae capsular polysaccharides 14 and 19F were measured in cord blood samples from 425 neonates, to determine which antibody subclass was most strongly associated with otitis media (OM) during the first 6 months of life (early OM). Early OM was significantly associated with type 14 IgG1 antibody in the lowest antibody quartile (P=.055) but not with type 19F IgG1 antibody or with either IgG2 antibody. IgG1 and IgG2 antibodies were significantly intercorrelated for type 14 (r=.52, P<.001) and type 19F (r=.38, P<.001). Multivariate analysis revealed that having type 14 IgG1 antibody in the lowest quartile, child care attendance, and sibling and maternal OM history were independent risk factors for early OM. Although type-specific pneumococcal IgG2 antibody concentrations were significantly higher than IgG1 concentrations, IgG2 antibodies apparently are not protective against OM during early infancy.

An analytical model applied to a multicenter pneumococcal enzyme-linked immunosorbent assay study.

Pneumococcal conjugate vaccines will eventually be licensed after favorable results from phase III efficacy trials. After licensure of a conjugate vaccine for invasive pneumococcal disease in infants, new conjugate vaccines will likely be licensed primarily on the basis of immunogenicity data rather than clinical efficacy. Analytical methods must therefore be developed, evaluated, and validated to compare immunogenicity results accurately within and between laboratories for different vaccines. At present no analytical technique is uniformly accepted and used in vaccine evaluation studies to determine the acceptable level of agreement between a laboratory result and the assigned value for a given serum sample. This multicenter study describes the magnitude of agreement among 12 laboratories quantifying an identical series of 48 pneumococcal serum specimens from 24 individuals (quality-control sera) by a consensus immunoglobulin G (IgG) enzyme-linked immunosorbent assay (ELISA) developed for this study. After provisional or trial antibody concentrations were assigned to the quality-control serum samples for this study, four methods for comparison of a series of laboratory-determined values with the assigned concentrations were evaluated. The percent error between assigned values and laboratory-determined concentrations proved to be the most informative of the four methods. We present guidelines that a laboratory may follow to analyze a series of quality-control sera to determine if it can reproduce the assigned antibody concentrations within an acceptable level of tolerance. While this study focused on a pneumococcal IgG ELISA, the methods that we describe are easily generalizable to other immunological assays.

Otitis media prevention: non-vaccine prophylaxis.

Preventing recurrent acute otitis media (AOM) is a goal of child health care. The objective is to reduce the frequency of pain and fever, shorten the duration of hearing loss, reduce the costs of physician visits, surgery and drugs, reduce parent anxiety, and prevent long term sequelae. Preventive approaches include understanding individual and familial risk factors, avoidance of environmental risk factors, antibiotic drug prophylaxis, polyvalent pneumococcal vaccination, myringotomy with tympanostomy tubes, and adenoidectomy. Earlier and more aggressive treatment can be provided to infants at increased risk. Antibiotic prophylaxis is challenged by a relatively small benefit and emerging resistant bacteria. Tympanostomy tubes are beneficial in chronic otitis media with effusion (OME), but of less value in recurrent AOM absent chronic OME. Adenoidectomy after tympanostomy tube failure is supported by at least one clinical trial.

Acute otitis media: management and surveillance in an era of pneumococcal resistance. Drug-Resistant Streptococcus Pneumoniae Therapeutic Working Group.

Experts in the management of otitis media and the Drug-resistant Streptococcus pneumoniae Therapeutic Working Group were convened by the Centers for Disease Control and Prevention to respond to changes in antimicrobial susceptibility among pneumococci. The objective was to provide consensus recommendations for the management of acute otitis media (AOM) and for the surveillance of drug-resistant Streptococcus pneumoniae. After summarizing published and unpublished data from the scientific literature and the experience of the panel members, the group concluded that oral amoxicillin should remain the first-line antimicrobial agent for treating AOM. For patients with clinically defined treatment failure after 3 days of therapy, useful alternative agents include amoxicillin-clavulanate, cefuroxime axetil, and intramuscular ceftriaxone. The group also made recommendations to improve surveillance and to obtain antimicrobial susceptibility patterns for local geographic areas.

Prospective study of tympanic membrane retraction, hearing loss, and multifrequency tympanometry.

Tympanic membrane retraction is a significant sequela of OME and has been linked clinically to atelectasis, ossicular erosion, and cholesteatoma. We investigated important factors for prediction of tympanic membrane retraction in a prospective study of 112 children. After 4 to 6 years of follow-up, 12% of ears had pars tensa retraction without atrophy, and 28% had various degrees of retraction with atrophy. Mild pars flaccida retraction was present in 23%, and severe pars flaccida retraction was present in 12%. Retraction severity was related to hearing level and multifrequency tympanometry. Three factors were significantly related to retraction severity: type of tube, male sex, and percent of visits in the second year with abnormal tympanograms. This study shows that type of tube was the most important factor in long-term outcome after tympanostomy tube treatment of OME.

Update of microbial problems in pediatric otorhinolaryngology: plenary session.

The bacteria to consider in upper respiratory tract infections in children are pneumococci, Haemophilus influenzae, group A streptococci and Moraxella catarrhalis.

Epidemiology of otitis media onset by six months of age.

OBJECTIVE: Although early otitis media (OM) onset predicts later recurrent and chronic OM, little research has been directed at illuminating the role of prenatal exposures in early OM. This prospective study examined prenatal, innate, and early environmental exposures associated with acute otitis media (AOM) onset and recurrent OM (ROM) by age 6 months. DESIGN AND METHODS: Prospective study of 596 infants from a health maintenance organization followed from birth to 6 months. Mothers completed monthly forms on prenatal exposures (diet, medications, and illnesses) and infant risk factors (eg, smoke exposure and child care) during pregnancy and until infants were 6 months old. Urine samples were collected when infants were 2 months of age and analyzed for cotinine and creatinine. Physicians and nurse practitioners examined infants at each clinic visit and completed standard ear examination forms. RESULTS: Thirty-nine percent had an episode of AOM and 20% had ROM by age 6 months. Using Cox's regression models to control for confounding, respiratory tract infection (relative risk [RR] 7.5), day care (RR 1. 7), >1 sibling (RR 1.4), maternal, paternal, and sibling OM history (RR 1.6, 1.5, and 1.7, respectively) were significantly related to early OM onset. ROM was related to respiratory tract infection (RR 9. 5), day care (RR 1.9), conjunctivitis (RR 2.0), maternal OM history (RR 1.9), and birth in the fall (RR 2.6). Among prenatal exposures, only high prenatal dietary vitamin C intake was significantly inversely related to early AOM with univariate but not multivariate analysis. CONCLUSION: Prenatal factors were not linked to early AOM onset with multivariate analysis, but environmental and innate factors play an important role in early AOM onset. Strategies to reduce exposure to environmental variables could reduce rates of early AOM, which could potentially result in declining rates of ROM and chronic OME.

Otitis media: the chinchilla model.

Streptococcus pneumoniae infection and disease have been modeled in several animal species including infant and adult mice, infant and adult rats, infant Rhesus monkeys, and adolescent and adult chinchillas. Most are models of sepsis arising from intravenous or intraperitoneal inoculation of bacteria, and a few were designed to study disease arising from intranasal infection. Chinchillas provide the only animal model of middle ear pneumococcal infection in which the disease can be produced by very small inocula injected into the middle ear (ME) or intranasally, and in which the disease remains localized to the ME in most cases. This model, developed at the University of Minnesota in 1975, has been used to study pneumococcal pathogenesis at a mucosal site, immunogenicity and efficacy of pneumococcal capsular polysaccharide (PS) vaccine antigens, and the kinetics and efficacy of antimicrobial drugs. Pathogenesis experiments in the chinchilla model have revealed variation in ME virulence among different pneumococcal serotypes, enhancement of ME infection during concurrent intranasal influenza A virus infections, and natural resolution of pneumococcal otitis media (OM) without intervention. Research has explored the relative contribution of pneumococcal and host products to ME inflammation. Pneumococcal cell wall components and pneumolysin have been studied in the model. Host inflammatory responses studied in the chinchilla ME include polymorphonuclear leukocyte oxidative products, hydrolytic enzymes, cytokine and eicosanoid metabolites, and ME epithelial cell adhesion and mucous glycoprotein production. Both clinical (tympanic membrane appearance) and histopathology (ME, Eustachian tube, inner ear) endpoints can be quantified. Immunologic and inflammatory studies have been facilitated by the production of affinity-purified antichinchilla immunoglobulin G (IgG), IgM, and secretory IgA polyclonal antibody reagents, and the identification of cross-reactivity between human and chinchilla cytokines, and between guinea pig and chinchilla C3. Alteration of ME mucosa by pneumococcal neuraminidase and alteration of ME epithelial cell (MEEC) surface carbohydrates during intranasal pneumococcal infection have been demonstrated. Pathogenesis studies have been aided by cultured chinchilla MEEC systems, in which the ability of platelet activating factor and interleukin (IL)-1 beta to stimulate epithelial mucous glycoprotein synthesis has recently been demonstrated. Because chronic OM with effusion is characterized by presence of large amounts of mucous glycoprotein in the ME, pneumococcus may have an important role in both acute and chronic ME disease. Both unconjugated PS and PS-protein-conjugated vaccines are immunogenic after intramuscular administration without adjuvant in chinchillas. Passive protection studies with human hyperimmune immunoglobulin demonstrated that anti-PS IgG alone is capable of protecting the chinchilla ME from direct ME challenge with pneumococci. Active PS immunization studies demonstrated protection following direct ME and intranasal pneumococcal challenge with and without concurrent influenza A virus infection. An attenuated influenza A virus vaccine also showed protection for pneumococcal OM. Antimicrobial treatment of acute OM has been based almost exclusively on empirical drug use and clinical trials without a foundation of ME pharmacokinetics. Studies in the chinchilla model have started to bring a rational basis to drug selection and dosing. Microassays have been developed using high-pressure liquid chromatography for many relevant drugs. Studies have explored the in vivo ME response in pneumococcal OM to antimicrobial drugs at supra- and sub-minimum inhibitory concentration (MIC), the effect of concurrent influenza A virus infection on ME drug penetration, and the effect of treatment on sensorineural hearing loss produced by pneumococcal OM.

Pneumococcus activation of the 5-lipoxygenase pathway and production of glycoproteins in the middle ear of rats.

Pneumococcal otitis media is associated with the production of potent inflammatory mediators (leukotrienes), but the mechanism by which pneumococcus induces production of leukotrienes in the middle ear is poorly understood. In this study, up-regulation of 2 genes that govern the lipoxygenase pathway, cPLA2 and 5-LOX, was observed in rats following inoculation of pneumococcus into the middle ear cavity. Expression of cPLA2 was low, and 5-LOX gene expression was not detected in control animals. Up-regulation of cPLA2 and 5-LOX in middle ear epithelial cells was accompanied by an increase of high-molecular-weight glycoproteins in middle ear fluid and cells. These findings suggest that pneumococcus activates the lipoxygenase pathway by up-regulating expression of the cPLA2 and 5-LOX genes. This, in turn, may stimulate synthesis and secretion of high-molecular-weight glycoproteins that facilitate production of fluid in the middle ear cleft.

Middle ear fluid cytokine and inflammatory cell kinetics in the chinchilla otitis media model.

Streptococcus pneumoniae is the most frequent microbe causing middle ear infection. The pathophysiology of pneumococcal otitis media has been characterized by measurement of local inflammatory mediators such as inflammatory cells, lysozyme, oxidative metabolic products, and inflammatory cytokines. The role of cytokines in bacterial infection has been elucidated with animal models, and interleukin (IL)-1beta, IL-6, and IL-8 and tumor necrosis factor alpha (TNF-alpha) are recognized as being important local mediators in acute inflammation. We characterized middle ear inflammatory responses in the chinchilla otitis media model after injecting a very small number of viable pneumococci into the middle ear, similar to the natural course of infection. Middle ear fluid (MEF) concentrations of IL-1beta, IL-6, IL-8, and TNF-alpha were measured by using anti-human cytokine enzyme-linked immunosorbent assay reagents. IL-1beta showed the earliest peak, at 6 h after inoculation, whereas IL-6, IL-8, and TNF-alpha concentrations were increasing 72 h after pneumococcal inoculation. IL-6, IL-8, and TNF-alpha but not IL-1beta concentrations correlated significantly with total inflammatory cell numbers in MEF, and all four cytokines correlated significantly with MEF neutrophil concentration. Several intercytokine correlations were significant. Cytokines, therefore, participate in the early middle ear inflammatory response to S. pneumoniae.

Chronic otitis media with effusion.

Chronic OME, which arises from a complex series of inflammatory events in the middle ear, affects approximately 5% to 30% of children. The mean duration of MEE is 16 to 20 weeks during the first 2 years of life. This condition is diagnosed best with pneumatic otoscopy and tympanometry. The risk of chronic OME is increased by environmental factors and characteristics of the child, including disease history. Approximately 70% of MEE are culture-positive, with approximately 50% of these yielding S pneumoniae, H influenzae, or M catarrhalis. However, antibiotic treatment of acute otitis media and OME has only a minimal effect on the long-term resolution of MEE. Research has shown that 70% of children who have chronic OME suffer mild-to-moderate hearing loss, so a child who has bilateral MEE for 3 months should undergo hearing evaluation. If the child has hearing impairment, referral to an otolaryngologist for myringotomy and tympanostomy tube insertion is a treatment option that the AHCPR recommends after 4 months of effusion with hearing loss. Sequelae of chronic OME include deficient expressive language and poorer attention skills due to the temporary hearing loss associated with OME, high-frequency sensorineural hearing loss, tympanic membrane atrophy, perforation, retraction, atelectasis, and cholesteatoma.

Acute otitis media: management and surveillance in an era of pneumococcal resistance--a report from the Drug-resistant Streptococcus pneumoniae Therapeutic Working Group.

OBJECTIVE: To provide recommendations [corrected] for the management of acute otitis media (AOM) and the surveillance of drug-resistant Streptococcus pneumoniae (DRSP). Five questions were addressed: (1) Can amoxicillin remain the best initial antimicrobial agent for treating AOM in the current period of increasing prevalence of DRSP? (2) What are suitable alternative agents for use if amoxicillin fails? (3) Should empiric treatment of AOM vary by geographic region? (4) Where can clinicians learn about resistance patterns in their patient populations? (5) What modifications to laboratory surveillance would improve the utility of the information for clinicians treating AOM? PARTICIPANTS: Experts in the management of otitis media and the DRSP Therapeutic Working Group. This group was convened by the CDC to respond to changes in antimicrobial susceptibility among pneumococci and includes clinicians, academicians and public health practitioners. EVIDENCE: Published and unpublished data summarized from the scientific literature and experience from the experts present. PROCESS: [corrected] After group presentations and review of background materials, subgroup chairs prepared draft responses to the five questions, discussed the responses as a group and edited those responses [corrected]. CONCLUSIONS: Oral amoxicillin should remain the first line antimicrobial agent for treating AOM. In view of the increasing prevalence of DRSP, the safety of amoxicillin at higher than standard dosages and evidence that higher dosages of amoxicillin can achieve effective middle ear fluid concentrations, an increase in the dosage used for empiric treatment from 40 to 45 mg/kg/day to 80 to 90 mg/kg/day is recommended. For patients with clinically defined treatment failure after 3 days of therapy, useful alternative agents include oral amoxicillin-clavulanate, cefuroxime axetil and intramuscular ceftriaxone. Many of the 13 other Food and Drug Administration-approved otitis media drugs lack good evidence for efficacy against DRSP. Currently local surveillance data for pneumococcal resistance that are relevant for the clinical management of AOM are not available from most areas in the United States. Recommendations to improve surveillance include establishing criteria for setting susceptibility breakpoints for clinically appropriate antimicrobials to ensure relevance for treating AOM, testing middle ear fluid or nasal swab isolates in addition to sterile site isolates and testing of drugs that are useful in treating AOM. The management of otitis media has entered a new era with the development of DRSP. These recommendations are intended to provide a framework for appropriate clinical and public health responses to this problem.

Relationships between otitis media sequelae and age.

OBJECTIVES: To explore relationships between age and sequelae in two groups of children treated with tympanostomy tubes for chronic otitis media with effusion (OME). STUDY DESIGN: Cross-sectional study of sequelae among children, adolescents, and adults at 4 years and 9 to 23 years after tympanostomy tube treatment. METHODS: Group I was examined with otomicroscopy, tympanometry, and audiometry two to four times a year as part of a prospective study, and they were evaluated 4 years after initial tube treatment for this study. Group II received tubes while participating in a chronic OME study, but participants were not followed prospectively after treatment. Nine to 23 years after tube treatment, they were examined with otomicroscopy, tympanometry, and hearing screening. RESULTS: Among the 5- to 28- year-old subjects, cholesteatoma (< or = 1%) and perforation (< or = 2%) were rare. In Group I, tympanosclerosis increased with age (P < .01), and OME (flat tympanograms) decreased with age in Group II (P < .01). The older cohort was more likely to have severe retractions (18% vs. 4%, P = .02), hearing loss (21% vs. 10%, P < .01), and severe atrophy (24% vs. 0%, P < .01) than the younger cohort, but they were less likely to have flat tympanograms (2% vs. 12%, P < .01). CONCLUSIONS: Although OME became less prevalent with age, important sequelae (severe atrophy, severe tympanic membrane retraction, hearing loss, cholesteatoma, and chronic perforation) may develop in children with chronic OME as they become adolescents and young adults. Long-term prospective studies are important in defining the progression of sequelae in these children.

Management of otitis media using Agency for Health Care Policy and Research guidelines. The Agency for Health Care Policy and Research.

Increased costs of managing otitis media and its complications may result from delays in diagnosis and treatment. The Agency for Health Care Policy and Research developed guidelines to assist in the management of chronic otitis media with effusion. We examined the medical care adherence to Agency for Health Care Policy and Research guidelines in 59 consecutive patients referred because of chronic otitis media with effusion and recurrent acute otitis media. Patient history and examination data were collected prospectively. In the group with chronic otitis media with effusion, the rate of adherence to Agency for Health Care Policy and Research guidelines was 0%; in those with recurrent acute otitis media, adherence was 5%. Delayed referral occurred in 34% of patients; 25% of patients were referred early. The average duration of effusion in patients with chronic otitis media with effusion was 5.2 months; the duration of recurrent acute otitis media immediately before referral was 9.3 months. Eighteen patients (47%) in the chronic otitis media with effusion group had a history of recurrent chronic otitis media with effusion spanning an average of 22.7 months. On referral, hearing loss was discovered in 92% of all patients, and in 69% the tympanogram was flat. The complication and sequelae rate was 49.1%, and speech delay was the most frequent at 16.9%. We conclude that in our study patients there is a significant referral delay, long history of chronic otitis media with effusion in patients before referral, high rate of hearing loss, and high complication rate. Continued efforts should be directed toward improving education of all clinicians so that diagnostic tools and timely otolaryngologic referral are better used.

Amoxicillin middle ear fluid penetration and pharmacokinetics in children with acute otitis media.

BACKGROUND: Acute otitis media (AOM) is a common childhood infectious disease. The efficacy of antibiotic dosing regimens is usually assessed by antibiotic plasma pharmacokinetics or middle ear fluid (MEF) concentration at one or two time points. Viral coinfection in AOM reduced antibacterial efficacy of antibiotics. OBJECTIVE: To determine amoxicillin MEF penetration and pharmacokinetics in bacterial and combined bacterial and viral AOM. METHODS: Thirty-four children with AOM were enrolled, and MEF was collected by tympanocentesis for bacterial culture and viral studies. Nasal wash and venous blood were also obtained for viral culture and serologic studies, respectively. Subjects were treated with amoxicillin 40 mg/kg/day orally, divided in equal doses every 8 h. During the second visit (48 to 72 h later) the subjects, with the regular morning amoxicillin dose withheld, were given an oral amoxicillin dose of 25 mg/kg. Thereafter two blood samples and one MEF sample by tympanocentesis were collected from each child at selected times between 0.5 and 4.0 h after dosing for bacterial and viral studies and amoxicillin concentration determination by high performance liquid chromatography. RESULTS: Eleven (37%) children had only bacterial infection, 6 (20%) had viral infection only, 6 (20%) had both bacterial and viral infections and in 7 (23%) neither bacterial nor viral pathogens were recovered. MEF bacterial culture was positive in 23 of 40 ears (57.5%) before treatment with amoxicillin (40 mg/kg/day) and was still positive in 4 of 38 ears (10.5%) after 2 to 3 days of treatment. Amoxicillin plasma concentration reached its peak at 1.0 to 1.5 h after a 25-mg/kg oral dose. The estimated MEF concentration peak occurred 3.0 h after the dose with MEF concentrations ranging from undetectable to 20.6 microg/ml and a mean of approximately 9.5 microg/ml. Geometric mean amoxicillin concentrations were lowest in virus-infected children (2.7 microg/ml), nearly the same in culture-negative samples from children without viral infection (2.9 microg/ml), higher in children with combined bacterial and viral infection (4.1 microg/ml) and highest in children with bacterial-only infection (5.7 microg/ml). CONCLUSIONS: MEF amoxicillin penetration tended to be lower in children with viral infection. The current amoxicillin dosing recommendation of 40 mg/kg/day in three divided dose is inadequate to effectively eradicate resistant Streptococcus pneumoniae, particularly during viral coinfection. A dosing regimen of 75 to 90 mg/kg/day is recommended for AOM.