Optical absorbance of the tympanic membrane in rat and human samples.

Chronic infections are often connected to biofilm formation. In presence of implants, this can lead to loss of the implant. Systemic or local application of drugs is relatively ineffective in case of biofilm formation. One technique to provide antibacterial properties on demand is the antibacterial photodynamic therapy (aPDT). Using this technique, these properties can be "switched on" by light illumination. In the middle ear with the semitransparent tympanic membrane, it might be possible in future to activate the antibacterial effect without opening the membrane. Therefore, we investigated the optical absorbance spectra of the tympanic membrane. Optical absorbance spectra were measured in ex vivo preparations from neonatal and adult rats with the membrane still being attached to the surrounding bony ring and four human samples. After performing area scans, the spot with the lowest absorbance being surrounded by a ring like structure with higher absorbance was chosen as region of interest for scanning wavelengths between 300 and 900 nm. Absorbance is generally higher at lower wavelengths with a local absorbance maximum at 420 nm and a weak second maximum with two neighbouring peaks at 540 / 580 nm and is significantly higher in adult rats compared to neonatal rats where about 10% of light was transmitted. The human samples show similar characteristics with a little higher absorbance. For activation of aPDT through the tympanic membrane, larger wavelengths are more promising. Whether the amount of light transmitted through the membrane would be sufficient to induce aPDT remains to be tested in further experiments.

Longitudinal optical coherence tomography to visualize the in vivo response of middle ear biofilms to antibiotic therapy.

Studying the impact of antibiotic treatment on otitis media (OM), the leading cause of primary care office visits during childhood, is critical to develop appropriate treatment strategies. Tracking dynamic middle ear conditions during antibiotic treatment is not readily applicable in patients, due to the limited diagnostic techniques available to detect the smaller amount and variation of middle ear effusion (MEE) and middle ear bacterial biofilm, responsible for chronic and recurrent OM. To overcome these challenges, a handheld optical coherence tomography (OCT) system has been developed to monitor in vivo response of biofilms and MEEs in the OM-induced chinchilla model, the standard model for human OM. As a result, the formation of MEE as well as biofilm adherent to the tympanic membrane (TM) was longitudinally assessed as OM developed. Various types of MEEs and biofilms in the chinchilla model were identified, which showed comparable features as those in humans. Furthermore, the effect of antibiotics on the biofilm as well as the amount and type of MEEs was investigated with low-dose and high-dose treatment (ceftriaxone). The capability of OCT to non-invasively track and examine middle ear conditions is highly beneficial for therapeutic OM studies and will lead to improved management of OM in patients.

Bacterial causes of otitis media with spontaneous perforation of the tympanic membrane in the era of 13 valent pneumococcal conjugate vaccine.

After pneumococcal conjugate vaccine (PCV) implementation, the number of acute otitis media (AOM) episodes has decreased, but AOM still remains among the most common diagnoses in childhood. From 2% to 17% of cases of AOM feature spontaneous perforation of the tympanic membrane (SPTM). The aim of this study was to describe the bacteriological causes of SPTM 5 to 8 years years after PCV13 implementation, in 2010. From 2015 to 2018, children with SPTM were prospectively enrolled by 41 pediatricians. Middle ear fluid was obtained by sampling spontaneous discharge. Among the 470 children with SPTM (median age 20.8 months), no otopathogen was isolated for 251 (53.4% [95% CI 48.8%;58.0%]): 47.1% of infants and toddlers, 68.3% older children (p<0.001). Among children with isolated bacterial otopathogens (n = 219), non-typable Haemophilus influenzae (NTHi) was the most frequent otopathogen isolated (n = 106, 48.4% [95% CI 41.6%;55.2%]), followed by Streptoccocus pyogenes (group A streptococcus [GAS]) (n = 76, 34.7% [95% CI 28.4%;41.4%]) and Streptococcus pneumoniae (Sp) (n = 61, 27.9% [95% Ci 22.0%;34.3%]). NTHi was frequently isolated in infants and toddlers (53.1%), whereas the main otopathogen in older children was GAS (52.3%). In cases of co-infection with at least two otopathogens (16.9%, n = 37/219), NTHi was frequently involved (78.4%, n = 29/37). When Sp was isolated, PCV13 serotypes accounted for 32.1% of cases, with serotype 3 the main serotype (16.1%). Among Sp strains, 29.5% were penicillin-intermediate and among NTHi strains, 16.0% were ß-lactamase-producers. More than 5 years after PCV13 implementation, the leading bacterial species recovered from AOM with SPTM was NTHi for infants and toddlers and GAS for older children. In both age groups, Sp was the third most frequent pathogen and vaccine serotypes still played an important role. No resistant Sp strains were isolated, and the frequency of ß-lactamase-producing NTHi did not exceed 16%.

Direct Analysis of Pathogenic Structures Affixed to the Tympanic Membrane during Chronic Otitis Media.

Objective To characterize otitis media-associated structures affixed to the mucosal surface of the tympanic membrane (TM) in vivo and in surgically recovered in vitro samples. Study Design Prospective case series without comparison. Setting Outpatient surgical care center. Subjects and Methods Forty pediatric subjects scheduled for tympanostomy tube placement surgery were imaged intraoperatively under general anesthesia. Postmyringotomy, a portable optical coherence tomography (OCT) imaging system assessed for the presence of any biofilm affixed to the mucosal surface of the TM. Samples of suspected microbial infection-related structures were collected through the myringotomy incision. The sampled site was subsequently reimaged with OCT to confirm collection from the original image site on the TM. In vitro analysis based on confocal laser scanning microscope (CLSM) images of fluorescence in situ hybridization-tagged samples and polymerase chain reaction (PCR) provided microbiological characterization and verification of biofilm activity. Results OCT imaging was achieved for 38 of 40 subjects (95%). Images from 38 of 38 (100%) of subjects observed with OCT showed the presence of additional microbial infection-related structures. Thirty-four samples were collected from these 38 subjects. CLSM images provided evidence of clustered bacteria in 32 of 33 (97%) of samples. PCR detected the presence of active bacterial DNA signatures in 20 of 31 (65%) of samples. Conclusion PCR and CLSM analysis of fluorescence in situ hybridization-stained samples validates the presence of active bacteria that have formed into a middle ear biofilm that extends across the mucosal layer of the TM. OCT can rapidly and noninvasively identify middle ear biofilms in subjects with severe and persistent cases of otitis media.

Factors affecting sound energy absorbance in acute otitis media model of chinchilla.

Acute otitis media (AOM) is a rapid-onset infection of the middle ear which results in middle ear pressure (MEP), middle ear effusion (MEE), and structural changes in middle ear tissues. Previous studies from our laboratory have identified that MEP, MEE, and middle ear structural changes are three factors affecting tympanic membrane (TM) mobility and hearing levels (Guan et al., 2014, 2013). Sound energy reflectance or absorbance (EA) is a diagnostic tool increasingly used in clinical settings for the identification of middle ear diseases. However, it is unclear whether EA can differentiate these three factors in an AOM ear. Here we report wideband EA measurements in the AOM model of chinchilla at three experimental stages: unopened, pressure released, and effusion removed. These correspond to the combined and individual effects of the three factors on sound energy transmission. AOM was produced by transbullar injection of Haemophilus influenzae in two treatment groups: 4 days (4D) and 8 days (8D) post inoculation. These time points represent the relatively early and later phase of AOM. In each group of chinchillas, EA at 250-8000 Hz was measured using a wideband tympanometer at three experimental stages. Results show that the effects of MEP, MEE, and tissue structural changes over the frequency range varied with the disease time course. MEP was the primary contributor to reduction of EA in 4D AOM ears and had a smaller effect in 8D ears. MEE reduced the EA at 6-8 kHz in 4D ears and 2-8 kHz in 8D ears and was responsible for the EA peak in both 4D and 8D ears. The residual EA loss due to structural changes was observed over the frequency range in 8D ears and only at high frequencies in 4D ears. The EA measurements were also compared with the published TM mobility loss in chinchilla AOM ears.

Role of Chlamydia pneumoniae and Helicobacteria pylori in the development of tympanosclerosis.

The etiology of tympanosclerosis (TS) is not known, but TS commonly develops secondary to acute and chronic otitis media (COM). Since calcification process in TS resembles that of atherosclerosis (AS), pathogens that are related to pathogenesis of AS may be involved in development of TS. This prospective and controlled study, performed at a tertiary referral center, investigated a possible relationship between the presence of Chlamydia (C.) pneumoniae and Helicobacter (H.) pylori and the development of a tympanosclerotic plaque. The presence of C. pneumoniae was examined in the surgical specimens of 62 patients (29 females and 33 males; age range 10-70 years, mean age 30.8 ± 13.3 years), including 30 patients with TS, 14 patients with cholesteatoma, and 18 patients with chronic suppurative otitis media (CSOM). The presence of H. pylori was examined in the surgical specimens of 88 patients (41 females and 47 males; age range 6-70 years, mean age 32.5 ± 14.8 years), including 35 patients with TS, 22 patients with cholesteatoma, 20 patients with CSOM, and 11 patients with otosclerosis. Tympanosclerotic plaques and control specimens from the cholesteatoma, polypoid mucosa, or mucosal portion of the perforations and stapes supra structure were examined for the presence of H. pylori and/or C. pneumoniae using real-time polymerase chain reaction analysis. The analysis demonstrated that specimens from the tympanosclerotic plaques and the other types of COM were all negative for C. pneumoniae and H. pylori. An association between C. pneumoniae or H. pylori infection and the development of TS or other types of COM could not be established.

Morphological changes in the tympanic membrane associated with Haemophilus influenzae-induced acute otitis media in the chinchilla.

OBJECTIVE: The tympanic membrane (TM) couples sound waves entering the outer ear canal to mechanical vibrations of the ossicular chain in the middle ear. During acute otitis media (AOM), dynamic structural changes in the TM can occur, which potentially affect sound transmission. It has remained unclear whether TM changes contribute significantly to the conductive hearing loss associated with human AOM. Studies that systematically and quantitatively assess the impact of morphological and mechanical characteristics of the TM on hearing in animal models of AOM have been few in number and lack detail. Our current study focused on the identification of quantitative morphological changes in the TM of the adult chinchilla. METHOD: AOM was produced by transbullar injection of the nontypeable (acapsular) Haemophilus influenzae strain 86-028NP into two treatment groups of chinchillas: one 4 days (4D) post bacterial challenge, and a second treatment group after 8 days (8D) post challenge. Structure and thickness were examined histologically at nine locations over the TM in untreated controls and in animals from both AOM treatment groups. RESULTS: TM thickness was found to have increased significantly (110-150%) at all measured locations of H. influenzae-infected ears when compared with uninfected (normal) TMs at 4D post bacterial challenge. Cellular proliferation and infiltration in the outer epithelial layer were primary contributors to this thickening. In ears infected for 8D, the TM was substantially thicker, a 200-300% increase from uninfected control values, due to edema and cell proliferation in both the outer and inner epithelial layers. In both 4D and 8D ears, thickening of the TM was more prominent in the superior-anterior quadrant. CONCLUSION: This study provides unequivocal structural evidence that significant TM thickness increases are associated with AOM induced by a well characterized H. influenzae human clinical isolate of low passage number. These and additional thickness data from early and later stages in middle ear infection will be used to derive the mechanical properties of the TM in a future study from our laboratory.

Factors affecting loss of tympanic membrane mobility in acute otitis media model of chinchilla.

Recently we reported that middle ear pressure (MEP), middle ear effusion (MEE), and ossicular changes each contribute to the loss of tympanic membrane (TM) mobility in a guinea pig model of acute otitis media (AOM) induced by Streptococcus pneumoniae (Guan and Gan, 2013). However, it is not clear how those factors vary along the course of the disease and whether those effects are reproducible in different species. In this study, a chinchilla AOM model was produced by transbullar injection of Haemophilus influenzae. Mobility of the TM at the umbo was measured by laser vibrometry in two treatment groups: 4 days (4D) and 8 days (8D) post inoculation. These time points represent relatively early and later phases of AOM. In each group, the vibration of the umbo was measured at three experimental stages: unopened, pressure-released, and effusion-removed ears. The effects of MEP and MEE and middle ear structural changes were quantified in each group by comparing the TM mobility at one stage with that of the previous stage. Our findings show that the factors affecting TM mobility do change with the disease time course. The MEP was the dominant contributor to reduction of TM mobility in 4D AOM ears, but showed little effect in 8D ears when MEE filled the tympanic cavity. MEE was the primary factor affecting TM mobility loss in 8D ears, but affected the 4D ears only at high frequencies. After the release of MEP and removal of MEE, residual loss of TM mobility was seen mainly at low frequencies in both 4D and 8D ears, and was associated with middle ear structural changes. Our findings establish that the factors contributing to TM mobility loss in the chinchilla ear were similar to those we reported previously for the guinea pig ears with AOM. Outcomes did not appear to differ between the two major bacterial species causing AOM in these animal models.

Could Helicobacter pylori play a role in the aetiopathogenesis of tympanosclerosis?

AIM: The aetiology of tympanosclerosis is not yet clear. This prospective, controlled, clinical study investigated the relationship between Helicobacter pylori and tympanosclerosis aetiology. MATERIALS AND METHODS: The study included 14 patients with tympanosclerosis and 26 with other forms of chronic otitis media. All patients underwent surgery for chronic otitis media. Mucosal biopsies were taken, and examined for H pylori using the Campylobacter-Like Organism (CLO) test. RESULTS: Tympanoplasty was performed in 29 patients (72.5 per cent), radical mastoidectomy in eight (20 per cent) and myringoplasty in three (7.5 per cent). The presence of H pylori was tested in all tympanosclerosis biopsies, but in only 26.9 per cent of biopsies from other forms of chronic otitis media. A statistically significant difference in H pylori presence was found (p ≤ 0.01). CONCLUSION: This study represents a preliminary investigation of the association between H pylori and tympanosclerosis development.

Treatment failure, recurrence, and antibiotic prescription rates for different acute otitis media treatment methods.

In an era of increasing antibiotic resistance, reliance on empirical antibiotic therapy for management of acute otitis media (AOM) will eventually become an untenable strategy. New research efforts are needed to identify effective treatment alternatives. In this retrospective analysis, we assessed AOM treatment failure (AOMTF), recurrent AOM (RAOM), and antibiotic prescription rates for groups of AOM episodes managed with either immediate antibiotics (ABX), tympanocentesis + observation (Tap + OBS), or tympanocentesis + immediate antibiotics (Tap + ABX). No significant differences in rates of AOMTF or RAOM were observed between any of the treatment groups. The 30-day rate of antibiotic prescriptions written for AOM was significantly lower in the TAP + OBS group (1 prescription per 3.8 episodes) than in the immediate antibiotic therapy groups (1.2 prescriptions per episode), P < .001. The comparative risk of AOMTF was slightly higher among AOM episodes caused by Streptococcus pneumoniae (OR 2.523; CI 1.070-5.944; P = 0.056). Neither the presence of other particular otopathogens, nor the presence of drug-resistant otopathogens, nor the choice of any particular AOM antibiotic therapy correlated with increased risk of either AOMTF or RAOM. The observed absence of significant differences in clinical outcomes between the various treatment groups in this study is consistent with findings published by other authors: alternative AOM therapies can help reduce reliance on antibiotics without inviting significant increases in rates of clinical failure.

Otogenic cerebellar abscess: a case report.

This case report describes the gradual deterioration of a healthy, highly functioning man who initially presented with a draining right ear. The patient's indolent neurologic decline and referral to an otologist ultimately led to the diagnosis and treatment of an otogenic cerebellar abscess, an increasingly rare intracranial complication of otitis media. We report this case to illustrate that severe complications of chronic otitis media still occur in the United States, to stress the importance of clinical suspicion in the postantibiotic era, and to review the literature regarding the most appropriate time to perform the otologic portion of the surgery.

[Peculiarities of clinical course of chronic purulent epitympanitis].

Wide application of antibiotics in bacterial infections have caused selection and prominent dissemination occurrence of antibioticoresistant strains of microorganisms. Etiological and pathogenetic peculiarities of chronic purulent epitympanitis (CHPE) were studied, rational curative scheme of 69 therapy patients was elaborated, which included sanation, the earpreserving operations on middle ear, performed for CHPE.

Aspergillus otomycosis in an immunocompromised patient.

Aspergillus niger, an opportunistic filamentous fungus, was identified as the cause of chronic unilateral otomycosis in a 55-year old, immunocompromised man who had been unresponsive to a variety of treatment regimens. The patient presented with intermittent otalgia and otorrhea and with a perforation of his left tympanic membrane. A niger was identified in a culture specimen obtained from the patient's left ear canal. In immunocompromised patients, it is important that the treatment of otomycosis be prompt and vigorous, to minimize the likelihood of hearing loss and invasive temporal bone infection.

Topical application of calcium channel blockers to reduce the progression of experimentally induced myringosclerosis and tympanosclerosis.

OBJECTIVES: This study aimed to evaluate the ability of topically applied calcium channel blockers (diltiazem) to reduce the progression of experimentally induced myringosclerosis and tympanosclerosis. STUDY DESIGN: Animal model. Experimental prospective study. METHODS: The study included 25 adult albino guinea pigs that were bilaterally myringotomized and inoculated with a suspension of Streptococcus pneumonia type 3. The right ears were treated with topical application of diltiazem, and the untreated left ears served as the control group. Otomicroscopy and remyringotomy were conducted every week. One animal was sacrificed after 1 week and the remaining at the end of 6 weeks. Temporal bones were dissected, and tympanic bullae were analyzed with light microscopy. RESULTS: The untreated control ears showed evidence of extensive myringosclerosis on otomicroscopy, and the ears treated with calcium channel blockers did as well although to a lesser degree. Under light microscopy, the lamina propria of both tympanic membranes and middle ear mucosae of the control group exhibited thicker (P < .1 and P < .05, respectively) and larger (P < .01 and P < .01, respectively) sclerotic tissue in comparison with the treatment group. CONCLUSION: The results suggest that calcium channel blockers had an influence in the prevention of tympanosclerosis.

[Establishment of tympanosclerosis by intratympanic injection of Staphylococcus aureus in guinea pigs].

OBJECTIVE: To establish a guinea pig model of tympanosclerosis and investigate the development and progression of tympanosclerosis in the tympanic membranes and middle ear mucosa. METHODS: Twenty-one healthy guinea pigs were subjected inoculation of 1x10(8)/L Staphylococcus aureus into the left middle ear cavities under general anaesthesia, with the right ears as the control, to establish models of chronic purulent otitis media. The animals were sacrificed by decapitation after 1, 3 and 6 months after model establishment for histological examination of the middle ear mucosa. RESULTS: The histological changes of tympanosclerosis occurred in some of the animals 3 months after model establishment, and the extent of calcium deposition and fibrosis across the mucosa were positively correlated with the duration of chronic purulent otitis media. CONCLUSION: Tympanosclerosis may occur in some guinea pigs after inoculation of Staphylococcus aureus into the middle ear cavity, and this model can be useful for study of tympanosclerosis.

Prediction of upper respiratory tract bacteria in acute otitis media.

CONCLUSIONS: Thorough otomicroscopical examination of the tympanic membrane in acute otitis media (AOM) might distinguish AOM episodes caused by different bacteria. It thus might be a way to select appropriate treatment for each patient without raising the number of dangerous complications. OBJECTIVES: The aim of this study was to see if it might be possible to predict the causative bacterium by judging the otomicroscopical appearance of the tympanic membrane in episodes of AOM. PATIENTS AND METHODS: The study was prospective. Patients suffering from non-perforated AOM were included. The tympanic membrane was photographed. A prediction of the causative bacterium was made and tympanocentesis was performed. Effusion from the middle ear and a nasopharyngeal swab were obtained for bacterial culturing. The causative bacteria were categorized into gram-positive (Streptococcus pneumoniae and S. pyogenes) or gram-negative (non-typable Haemophilus influenzae and Moraxella catarrhalis). RESULTS: A total of 82 patients were included in the study. A correct prediction was made in 47/63, a false prediction in 16/63 (kappa 0.48, p<0.001).

Otomicroscopic findings and systemic interleukin-6 levels in relation to etiologic agent during experimental acute otitis media.

The aim of the present study was to explore whether it was possible to differentiate the clinical course and the otomicroscopic appearance of acute otitis media (AOM) caused by common otitis pathogens in an animal model. Systemic interleukin (IL)-6 levels as early markers for bacterial AOM were also studied. Four groups of rats were inoculated with either Streptococcus pneumoniae, Streptococcus pyogenes, non-typeable Haemophilus influenzae or Moraxella catarrhalis. The animals were monitored by otomicroscopy, photos of the tympanic membrane, cultures and IL-6 detection in serum the following 4 days. The gram-positive S. pneumoniae and S. pyogenes induced severe AOM with opaque effusion behind the tympanic membrane, pronounced dilation of the vessels and spontaneous perforations. The gram-negative H. influenzae and M. catarrhalis induced a less severe infection with cloudy, sometimes foamy effusion, and no spontaneous perforations. With the otomicroscopic findings it was possible to distinguish between infections induced by gram-positive bacteria and gram-negative bacteria. Detection of interleukin-6 in serum appeared to be of limited use for all infections except the pneumococcal AOM, but this needs to be further investigated.

Pseudomonas aeruginosa infection in the hypoventilated middle ear: an experimental model.

CONCLUSION: This is a suitable model for studying different aspects of the pathophysiology of chronic suppurative otitis media. OBJECTIVE: To analyze the methodological features of an animal model of chronic suppurative otitis media induced by intratympanic inoculation of Pseudomonas aeruginosa. MATERIAL AND METHODS: Otitis media was induced by inoculation of P. aeruginosa through the inferior aspect of the rat bulla and cauterization of the Eustachian tube via a transpalatal approach. Inspection of the tympanic membrane, culturing of middle ear effusion and processing of the temporal bones for light microscopy were performed. RESULTS: Abnormal otomicroscopic findings and persistence of infection were correlated with the histopathological changes found in middle ear tissues.