FUT2 Variants Confer Susceptibility to Familial Otitis Media.

Non-secretor status due to homozygosity for the common FUT2 variant c.461G>A (p.Trp154∗) is associated with either risk for autoimmune diseases or protection against viral diarrhea and HIV. We determined the role of FUT2 in otitis media susceptibility by obtaining DNA samples from 609 multi-ethnic families and simplex case subjects with otitis media. Exome and Sanger sequencing, linkage analysis, and Fisher exact and transmission disequilibrium tests (TDT) were performed. The common FUT2 c.604C>T (p.Arg202∗) variant co-segregates with otitis media in a Filipino pedigree (LOD = 4.0). Additionally, a rare variant, c.412C>T (p.Arg138Cys), is associated with recurrent/chronic otitis media in European-American children (p = 1.2 × 10-5) and US trios (TDT p = 0.01). The c.461G>A (p.Trp154∗) variant was also over-transmitted in US trios (TDT p = 0.01) and was associated with shifts in middle ear microbiota composition (PERMANOVA p < 10-7) and increased biodiversity. When all missense and nonsense variants identified in multi-ethnic US trios with CADD > 20 were combined, FUT2 variants were over-transmitted in trios (TDT p = 0.001). Fut2 is transiently upregulated in mouse middle ear after inoculation with non-typeable Haemophilus influenzae. Four FUT2 variants-namely p.Ala104Val, p.Arg138Cys, p.Trp154∗, and p.Arg202∗-reduced A antigen in mutant-transfected COS-7 cells, while the nonsense variants also reduced FUT2 protein levels. Common and rare FUT2 variants confer susceptibility to otitis media, likely by modifying the middle ear microbiome through regulation of A antigen levels in epithelial cells. Our families demonstrate marked intra-familial genetic heterogeneity, suggesting that multiple combinations of common and rare variants plus environmental factors influence the individual otitis media phenotype as a complex trait.

A2ML1 and otitis media: novel variants, differential expression, and relevant pathways.

A genetic basis for otitis media is established, however, the role of rare variants in disease etiology is largely unknown. Previously a duplication variant within A2ML1 was identified as a significant risk factor for otitis media in an indigenous Filipino population and in US children. In this report exome and Sanger sequencing was performed using DNA samples from the indigenous Filipino population, Filipino cochlear implantees, US probands, Finnish, and Pakistani families with otitis media. Sixteen novel, damaging A2ML1 variants identified in otitis media patients were rare or low-frequency in population-matched controls. In the indigenous population, both gingivitis and A2ML1 variants including the known duplication variant and the novel splice variant c.4061 + 1 G>C were independently associated with otitis media. Sequencing of salivary RNA samples from indigenous Filipinos demonstrated lower A2ML1 expression according to the carriage of A2ML1 variants. Sequencing of additional salivary RNA samples from US patients with otitis media revealed differentially expressed genes that are highly correlated with A2ML1 expression levels. In particular, RND3 is upregulated in both A2ML1 variant carriers and high-A2ML1 expressors. These findings support a role for A2ML1 in keratinocyte differentiation within the middle ear as part of otitis media pathology and the potential application of ROCK inhibition in otitis media.

Otitis Media in Children with Down Syndrome Is Associated with Shifts in the Nasopharyngeal and Middle Ear Microbiotas.

Background: Otitis media (OM) is defined as middle ear (ME) inflammation that is usually due to infection. Globally, OM is a leading cause of hearing loss and is the most frequently diagnosed disease in young children. For OM, pediatric patients with Down syndrome (DS) demonstrate higher incidence rates, greater severity, and poorer outcomes. However, to date, no studies have investigated the bacterial profiles of children with DS and OM. Method: We aimed to determine if there are differences in composition of bacterial profiles or the relative abundance of individual taxa within the ME and nasopharyngeal (NP) microbiotas of pediatric OM patients with DS (n = 11) compared with those without DS (n = 84). We sequenced the 16S rRNA genes and analyzed the sequence data for diversity indices and relative abundance of individual taxa. Results: Individuals with DS demonstrated increased biodiversity in their ME and NP microbiotas. In children with OM, DS was associated with increased biodiversity and higher relative abundance of specific taxa in the ME. Conclusion: Our findings suggest that dysbioses in the NP of DS children contributes to their increased susceptibility to OM compared with controls. These findings suggest that DS influences regulation of the mucosal microbiota and contributes to OM pathology.

Pediatric cochlear implantation.

PURPOSE OF REVIEW: To summarize the current state of cochlear implantation in children. RECENT FINDINGS: Hearing loss profoundly impacts a child's ability to communicate and thereby affects educational and psychosocial development. The auditory cortex has sensitive periods during which it is maximally receptive to sound stimulus and consequent development. For this reason, there is impetus to implant children at a very young age, as a cochlear implant can provide auditory input during this critical window. Long-term follow-up of implanted children has shown that cochlear implants are able to provide substantial language, academic, and social benefit. The criteria for cochlear implantation have broadened, and promising outcomes are being seen in populations that were not previously considered for implantation. The best language acquisition outcomes are seen in recipients who were implanted at a younger age and are in an environment rich in oral communication. Bilateral implantation offers some acoustic advantages. SUMMARY: Cochlear implants provide sound perception to deaf children and can mitigate, to varying extents, the effects of sound deprivation on auditory development. On the basis of our understanding of brain development and language outcomes, a wider population of children are now candidates for implantation than previously considered.

The FUT2 Variant c.461G>A (p.Trp154*) Is Associated With Differentially Expressed Genes and Nasopharyngeal Microbiota Shifts in Patients With Otitis Media.

Otitis media (OM) is a leading cause of childhood hearing loss. Variants in FUT2, which encodes alpha-(1,2)-fucosyltransferase, were identified to increase susceptibility to OM, potentially through shifts in the middle ear (ME) or nasopharyngeal (NP) microbiotas as mediated by transcriptional changes. Greater knowledge of differences in relative abundance of otopathogens in carriers of pathogenic variants can help determine risk for OM in patients. In order to determine the downstream effects of FUT2 variation, we examined gene expression in relation to carriage of a common pathogenic FUT2 c.461G>A (p.Trp154*) variant using RNA-sequence data from saliva samples from 28 patients with OM. Differential gene expression was also examined in bulk mRNA and single-cell RNA-sequence data from wildtype mouse ME mucosa after inoculation with non-typeable Haemophilus influenzae (NTHi). In addition, microbiotas were profiled from ME and NP samples of 65 OM patients using 16S rRNA gene sequencing. In human carriers of the FUT2 variant, FN1, KMT2D, MUC16 and NBPF20 were downregulated while MTAP was upregulated. Post-infectious expression in the mouse ME recapitulated these transcriptional differences, with the exception of Fn1 upregulation after NTHi-inoculation. In the NP, Candidate Division TM7 was associated with wildtype genotype (FDR-adj-p=0.009). Overall, the FUT2 c.461G>A variant was associated with transcriptional changes in processes related to response to infection and with increased load of potential otopathogens in the ME and decreased commensals in the NP. These findings provide increased understanding of how FUT2 variants influence gene transcription and the mucosal microbiota, and thus contribute to the pathology of OM.

The role of CDHR3 in susceptibility to otitis media.

Otitis media (OM) is common in young children and can cause hearing loss and speech, language, and developmental delays. OM has high heritability; however, little is known about OM-related molecular and genetic processes. CDHR3 was previously identified as a locus for OM susceptibility, but to date, studies have focused on how the CDHR3 p.Cys529Tyr variant increases epithelial binding of rhinovirus-C and risk for lung or sinus pathology. In order to further delineate a role for CDHR3 in OM, we performed the following: exome sequencing using DNA samples from OM-affected individuals from 257 multi-ethnic families; Sanger sequencing, logistic regression and transmission disequilibrium tests for 407 US trios or probands with OM; 16S rRNA sequencing and analysis for middle ear and nasopharyngeal samples; and single-cell RNA sequencing and differential expression analyses for mouse middle ear. From exome sequence data, we identified a novel pathogenic CDHR3 splice variant that co-segregates with OM in US and Finnish families. Additionally, a frameshift and six missense rare or low-frequency variants were identified in Finnish probands. In US probands, the CDHR3 p.Cys529Tyr variant was associated with the absence of middle ear fluid at surgery and also with increased relative abundance of Lysobacter in the nasopharynx and Streptomyces in the middle ear. Consistent with published data on airway epithelial cells and our RNA-sequence data from human middle ear tissues, Cdhr3 expression is restricted to ciliated epithelial cells of the middle ear and is downregulated after acute OM. Overall, these findings suggest a critical role for CDHR3 in OM susceptibility. KEY MESSAGES: • Novel rare or low-frequency CDHR3 variants putatively confer risk for otitis media. • Pathogenic variant CDHR3 c.1653 + 3G > A was found in nine families with otitis media. • CDHR3 p.Cys529Tyr was associated with lack of effusion and bacterial otopathogens. • Cdhr3 expression was limited to ciliated epithelial cells in mouse middle ear. • Cdhr3 was downregulated 3 h after infection of mouse middle ear.

Behavioral Audiology Procedures in Children With Down Syndrome.

Purpose Normative data regarding behavioral audiologic testing procedures are based upon the general population and often do not apply to children with Down syndrome (DS). Testing children with DS can be challenging, and outcomes may be unreliable due to their different cognitive demands and delays. The aim of this study was to assess optimal audiologic testing procedures for specific age groups of children with DS. Method This study used a retrospective investigation of 273 children with DS (145 boys, 128 girls; average age at evaluation = 5.92 ± 4.74 years) who received an audiologic evaluation during 2013 as part of their medical care at a large pediatric hospital (satellite facilities included). Results Age ranges for the completion of audiometry procedures in children with DS are provided. Average age to reliably complete behavioral testing in children with DS was delayed by up to 30 months compared to typically developing children. The majority of children with DS achieved at least good-to-fair reliability for audiologic results starting at 16 months (85.7%) and two ear results at 6-10 years (76.1%). Though not statistically significant, the use of a two-tester assistant compared to a single tester appeared to be helpful in obtaining reliable results. Conclusion The results provide a guide to optimal audiologic test procedures for children with DS, as the standard audiologic guidelines for typically developing infants and children do not apply.

Novel Variants in Hearing Loss Genes and Associations With Audiometric Thresholds in a Multi-ethnic Cohort of US Patients With Cochlear Implants.

OBJECTIVES: To investigate novel variants in hearing loss genes and clinical factors affecting audiometric outcomes of cochlear implant (CI) patients. BACKGROUND: Approximately 50% of hearing loss has a genetic etiology, with certain genetic variants more prevalent in specific ethnic groups. Different variants and some clinical variables including inner ear malformations result in different prognoses or clinical outcomes after CI. METHODS: Medical and genetic testing records of pediatric CI patients were reviewed for clinical variables. Minor allele frequencies of variants were obtained from Genome Aggregation Database (gnomAD) and variants were classified for pathogenicity. Standard statistical testing was done using Fisher's exact, Wilcoxon, and Spearman correlation tests. RESULTS: Eighteen CI patients with genetic test results had pathogenic variants, including six patients with syndromic hearing loss and six patients with known GJB2 variants. Novel pathogenic variants were noted in CHD7, ADGRV1, and ARID1B, with variants in the latter two genes identified in Hispanic patients. Overall, carriage of genetic variants was associated with better pre-CI audiometric thresholds at 2000 Hz (p = 0.048). On the other hand, post-CI thresholds were significantly worse in patients with inner ear malformations, particularly in patients with atretic cochlear nerve canals. CONCLUSION: Four novel pathogenic variants were identified, which contributes to knowledge of allelic spectrum for hearing loss especially in Hispanic patients. In this cohort, carriage of pathogenic variants particularly of GJB2 variants was associated with better pre-CI audiometric thresholds, while patients with inner ear malformations had worse post-CI audiometric thresholds.

Multi-omic studies on missense PLG variants in families with otitis media.

Otitis media (OM), a very common disease in young children, can result in hearing loss. In order to potentially replicate previously reported associations between OM and PLG, exome and Sanger sequencing, RNA-sequencing of saliva and middle ear samples, 16S rRNA sequencing, molecular modeling, and statistical analyses including transmission disequilibrium tests (TDT) were performed in a multi-ethnic cohort of 718 families and simplex cases with OM. We identified four rare PLG variants c.112A > G (p.Lys38Glu), c.782G > A (p.Arg261His), c.1481C > T (p.Ala494Val) and c.2045 T > A (p.Ile682Asn), and one common variant c.1414G > A (p.Asp472Asn). However TDT analyses for these PLG variants did not demonstrate association with OM in 314 families. Additionally PLG expression is very low or absent in normal or diseased middle ear in mouse and human, and salivary expression and microbial α-diversity were non-significant in c.1414G > A (p.Asp472Asn) carriers. Based on molecular modeling, the novel rare variants particularly c.782G > A (p.Arg261His) and c.2045 T > A (p.Ile682Asn) were predicted to affect protein structure. Exploration of other potential disease mechanisms will help elucidate how PLG contributes to OM susceptibility in humans. Our results underline the importance of following up findings from genome-wide association through replication studies, preferably using multi-omic datasets.

Identification of Novel Genes and Biological Pathways That Overlap in Infectious and Nonallergic Diseases of the Upper and Lower Airways Using Network Analyses.

Previous genetic studies on susceptibility to otitis media and airway infections have focused on immune pathways acting within the local mucosal epithelium, and outside of allergic rhinitis and asthma, limited studies exist on the overlaps at the gene, pathway or network level between the upper and lower airways. In this report, we compared [1] pathways identified from network analysis using genes derived from published genome-wide family-based and association studies for otitis media, sinusitis, and lung phenotypes, to [2] pathways identified using differentially expressed genes from RNA-sequence data from lower airway, sinus, and middle ear tissues, in particular cholesteatoma tissue compared to middle ear mucosa. For otitis media, a large number of genes (n = 1,806) were identified as differentially expressed between cholesteatoma and middle ear mucosa, which in turn led to the identification of 68 pathways that are enriched in cholesteatoma. Two differentially expressed genes CR1 and SAA1 overlap in middle ear, sinus, and lower airway samples and are potentially novel genes for otitis media susceptibility. In addition, 56 genes were differentially expressed in both tissues from the middle ear and either sinus or lower airways. Pathways that are common in upper and lower airway diseases, whether from published DNA studies or from our RNA-sequencing analyses, include chromatin organization/remodeling, endocytosis, immune system process, protein folding, and viral process. Taken together, our findings from genetic susceptibility and differential tissue expression studies support the hypothesis that the unified airway theory wherein the upper and lower respiratory tracts act as an integrated unit also applies to infectious and nonallergic airway epithelial disease. Our results may be used as reference for identification of genes or pathways that are relevant to upper and lower airways, whether common across sites, or unique to each disease.

Pediatric vocal fold paralysis after cardiac surgery: rate of recovery and sequelae.

OBJECTIVE: To determine the rate of recovery of pediatric vocal fold paralysis (VFP) after cardiac surgery. STUDY DESIGN AND SETTING: Retrospective case series from January 2000 to 2005 at 4 tertiary care pediatric hospitals. RESULTS: A total of 109 children with VFP were identified. Of 80 patients with follow-up >3 months, 28 (35%) recovered vocal fold function with a median time to diagnosis of recovery of 6.6 months. Fifty-two (65%) patients had persistent vocal fold paralysis with a median follow-up time of 16.4 months. Twenty-five (45%) of 55 patients demonstrated aspiration or laryngeal penetration with modified barium swallow. Twenty-nine (27%) of the 109 patients underwent surgical intervention for their airway, feeding, or voice. CONCLUSIONS: Pediatric VFP is not an uncommon complication after cardiac surgery and can result in serious sequelae. This study demonstrates a 35% rate of recovery, 45% rate of aspiration, and 27% rate of complications that require surgical intervention.

Interrater and intrarater reliability in the evaluation of velopharyngeal insufficiency within a single institution.

OBJECTIVE: To explore the interrater and intrarater reliability in nasoendoscopic assessment of velopharyngeal (VP) function using the standardized reporting method described by Golding-Kushner within a single institution. DESIGN: Prospective blinded study. SETTING: Academic, tertiary care, pediatric hospital. PARTICIPANTS: Six health care providers (2 pediatric otolaryngology faculty members, 2 pediatric otolaryngology fellows, and 2 speech pathologists) independently rated 50 videotaped nasoendoscopy segments twice. The segments on the videotape were obtained in a clinical setting. MAIN OUTCOME MEASURES: The Golding-Kushner rating system was used to rate VP function. Raters described VP closure quantitatively by rating palatal and lateral pharyngeal wall movement for each segment. They also qualitatively described characteristics of the VP gap, rated gap size as none, small, medium, or large, and estimated the percentage gap size relative to the resting position. Reliability coefficients were calculated for the data sets. RESULTS: Fairly good interrater and intrarater reliability was seen in the quantitative measures. Faculty otolaryngologists rated segments more similarly to each other than did pediatric otolaryngology fellows, but intrarater reliability was similar for both the experienced and less experienced otolaryngologists. Less consistency was seen in the ratings of the speech pathologists. Raters tended to rate with less consistency when describing qualitative characteristics of the VP gap than when making quantitative measurements. CONCLUSIONS: The Golding-Kushner scale is a reasonably reliable tool for reporting nasoendoscopic findings at our institution. However, these data also indicate that there exists room for improvement and that rater training may increase reliability.

Partial promontory technique in stapedotomy cases with narrow niche.

OBJECTIVE: The objective of this study was to examine clinical and audiometric outcomes of a laser partial promontory technique in stapedotomy cases with a narrow oval window niche. STUDY DESIGN: We conducted a retrospective chart review. SETTING: This study was conducted at a tertiary referral center. PATIENTS: We studied 59 patients who underwent a partial promontory technique with stapedotomy between 1994 and 2000. Seventy-two patients who underwent primary stapedotomy without promontory technique served as a control group. METHODS: Preoperative and postoperative audiometric results were obtained for 59 patients undergoing laser stapedotomy with a narrow oval window niche. The partial promontory removal was performed with a KTP laser. Results were compared with 72 primary laser stapedotomy cases without the promontory technique within the same time period and analyzed using paired Student t test. RESULTS: Ninety percent of the partial promontory cases were successful (air-bone gap [ABG] <10 dB). The mean postoperative ABG was 5.1 dB, which was comparable to the non-promontory cases (p = 0.7). The mean change in postoperative bone conduction was also comparable (p = 0.98). There were no cases of sensorineural hearing loss. An overhanging facial nerve was present in 32% of the narrow niche cases and a dehiscent facial nerve was encountered in 17% of these cases. CONCLUSIONS: Partial laser removal of the promontory as an adjunct to laser stapedotomy cases with a narrow oval window niche is a safe, effective technique with comparable results to primary laser stapedotomy.

Prevention and correction of nasal tip bossae in rhinoplasty.

OBJECTIVE: To describe our experiences with nasal tip bossae, suggest a standard nomenclature, discuss causative factors, and provide a comprehensive, analytic approach to the prevention and correction of bossae. BACKGROUND: Nasal tip bossae are knoblike protuberances of the alar cartilages that can arise after rhinoplasty. Early bossae are due to uncorrected or inadvertently created asymmetries, while late bossae are due to fibrosis and scar contracture acting on a weakened or unreconstituted cartilaginous framework. Numerous techniques may be used to prevent and treat bossae; however, we found no article in the existing literature that presents an in-depth, analytic description of management techniques. METHODS: We analyzed the predisposing factors and techniques leading to bossa formation and studied principles of prevention and correction. All rhinoplasty cases that presented for revision from 1985 through 2000 were reviewed for bossae formation via internal computer search. Previous operative records for rhinoplasty cases were examined when available. Intraoperative notes and photgraphs of the revision surgery were examined. RESULTS: Etiologies for bossae were consistently found, and successful treatment modalities were noted. CONCLUSIONS: Nasal tip bossae are most often due to dynamic forces acting on iatrogenic changes and/or weakness in the alar cartilages. By minimizing cartilage excision, reinforcing areas of weakness, avoiding asymmetry and irregularity, and maintaining alar integrity, formation of bossae may be prevented. The treatment of bossae must be individualized and can range from simple suture stabilization techniques to complex domal cartilage replacement grafts, depending on the observed defect.

The need for long-term audiologic follow-up of neonatal intensive care unit (NICU) graduates.

OBJECTIVE: To evaluate the adequacy of newborn hearing screening in the identification of hearing loss in post-neonatal intensive care unit (NICU) infants. METHODS: Eighty-two post-NICU infants who had initially passed automated auditory brainstem response (AABR) screening were studied prospectively between November 1997 and July 1999. Tympanometry and transient evoked otoacoustic emissions (TEOAE) were used to evaluate middle ear status and screen the hearing of subjects when they were seen routinely in the Mary L. Johnson Infant Development Clinic, where NICU graduates are followed at our institution. TEOAEs were not performed in subjects with abnormal tympanometry, defined as negative pressures greater than 200 daPa or flat tympanograms. RESULTS: Of the 82 subjects, 31 (37%) had abnormal tympanometry in at least one ear, with 24 (29%) exhibiting abnormal values bilaterally. Two subjects were identified with delayed-onset or previously undiagnosed sensorineural hearing loss. One had a history of persistent pulmonary hypertension (PPHN) and extracorporeal membrane oxygenation. The other infant had no risk factors for sensorineural hearing loss. CONCLUSIONS: Our data indicate that newborn hearing screening programs may not provide adequate vigilance for NICU graduates. The high incidence of abnormal middle ear status and the identification of delayed-onset hearing loss in an infant without known risk factors highlights the need for close audiologic and speech/language follow-up in the post-NICU population.

Clinicopathological characteristics of extramedullary acute megakaryoblastic leukemia (AMKL): report of a case with initial mastoid presentation and review of literature to compare extramedullary AMKL and non-AMKL cases.

Extramedullary acute megakaryoblastic leukemia (AMKL) is a rare neoplasm with a varied clinical presentation. AMKL with initial mastoid presentation has never been reported. The extreme rarity of mastoid AMKL, together with the tendency of extramedullary AMKL to mimic other small blue cell tumors, can create a diagnostic challenge. We report a case of AMKL that initially presented as a mastoid lesion and provide a comprehensive review and analysis that compares the characteristics of extramedullary AMKL and nonmegakaryoblastic acute myeloid leukemia (AML) in reported pediatric cases over the past 30 years. We found that patients with extramedullary AMKL were not only younger than patients without megakaryocytic differentiation but were also limited to those ≤ 2 years of age. In addition, girls predominated in both AMKL and AML MLL(+) groups compared with other types of AML (P  =  0.0366 and P  =  0.0082). Furthermore, we found that extramedullary AMKL was more likely to involve bone than AML MLL(+) (P < 0.0001) or other types of AML (P  =  0.0002). These findings suggest that extramedullary AMKL should be considered in the differential diagnosis of SBCT in children, especially in patients with mastoid or other bony lesions, those ≤ 2 years of age, and female patients.

Generation of consensus in the application of a rating scale to nasendoscopic assessment of velopharyngeal function.

OBJECTIVE: To generate consensus ratings of velopharyngeal function on nasendoscopy (NE) with the goal of creating a video instruction tool. METHODS: The American Society of Pediatric Otolaryngology Velopharyngeal Insufficiency Study Group convened to identify NE segments to be included in an instructional video. Of 24 segments reviewed, 11 were selected based on the quality of the examinations and spectrum of closure patterns. Participating otolaryngologists independently rated NE segments using the Golding-Kushner scale. The participants then convened and rated each of the NE segments as a group. Thirty-nine members of the American Society of Pediatric Otolaryngology met and agreed with the group ratings, creating a consensus standard. RESULTS: Individual scores for palate and lateral wall motion showed high variability, ranging from 0 to 6 points difference from the consensus. Variability was also seen for the following qualitative findings: the Passavant ridge, aberrant pulsations, and dorsal palatal notch. The individual ratings are presented graphically to demonstrate the range of individual responses as well as to compare responses to the consensus ratings. No further changes were made to the proposed consensus ratings when reviewed by the larger group. CONCLUSIONS: Rating of NE evaluations of velopharyngeal function was variable among a group of pediatric otolaryngologists experienced in treating velopharyngeal insufficiency. These results highlight the need to develop a standardized method of reporting NE findings for velopharyngeal insufficiency. Despite this, consensus ratings were achieved that will facilitate development of a video instruction tool.