Reliability of nasopharyngeal PCR for the detection of otopathogens in children with uncomplicated acute otitis media compared to culture.

Otopathogens in acute otitis media (AOM) have implications for care because the likelihood of resolution without antibiotics and optimal antibiotic agent varies by microorganism. We aimed to determine the sensitivity, specificity, positive predictive value, and negative predictive value of nasopharyngeal (NP) qualitative polymerase chain reaction (PCR) for common bacterial otopathogens in children with AOM compared to NP culture. NP flocked swabs collected from enrolled children aged 6 to 35 months with uncomplicated AOM in Denver, CO were tested by culture and multiplex PCR. The sensitivity and negative predictive value of PCR using culture as a reference were high (H. influenzae 93.3%, 98.0%; S. pneumoniae 94.2%, 95.1%; M. catarrhalis 92.3%, 86.4%); whereas the specificity and positive predictive value were lower and varied by organism (54.2%-84.1%, 55.1%-69.2%, respectively). PCR detected 1.5 times more organisms than culture. NP PCR has a high predictive value for excluding otopathogens compared to culture and warrants exploration as a diagnostic tool.

Innate Immunity in the Middle Ear Mucosa.

Otitis media (OM) encompasses a spectrum of clinical presentations ranging from the readily identifiable Acute OM (AOM), which is characterised by otalgia and fever, to chronic otitis media with effusion (COME) where impaired hearing due to middle ear effusion may be the only clinical symptom. Chronic suppurative OM (CSOM) presents as a more severe form of OM, involving perforation of the tympanic membrane. The pathogenesis of OM in these varied clinical presentations is unclear but activation of the innate inflammatory responses to viral and/or bacterial infection of the upper respiratory tract performs an integral role. This localised inflammatory response can persist even after pathogens are cleared from the middle ear, eustachian tubes and, in the case of respiratory viruses, even the nasal compartment. Children prone to OM may experience an over exuberant inflammatory response that underlies the development of chronic forms of OM and their sequelae, including hearing impairment. Treatments for chronic effusive forms of OM are limited, with current therapeutic guidelines recommending a "watch and wait" strategy rather than active treatment with antibiotics, corticosteroids or other anti-inflammatory drugs. Overall, there is a clear need for more targeted and effective treatments that either prevent or reduce the hyper-inflammatory response associated with chronic forms of OM. Improved treatment options rely upon an in-depth understanding of OM pathogenesis, particularly the role of the host innate immune response during acute OM. In this paper, we review the current literature regarding the innate immune response within the middle ear to bacterial and viral otopathogens alone, and as co-infections. This is an important consideration, as the role of respiratory viruses as primary pathogens in OM is not yet fully understood. Furthermore, increased reporting from PCR-based diagnostics, indicates that viral/bacterial co-infections in the middle ear are more common than bacterial infections alone. Increasingly, the mechanisms by which viral/bacterial co-infections may drive or maintain complex innate immune responses and inflammation during OM as a chronic response require investigation. Improved understanding of the pathogenesis of chronic OM, including host innate immune response within the middle ear is vital for development of improved diagnostic and treatment options for our children.

Nasopharyngeal colonization of otopathogens in South Indian children with acute otitis media - A case control pilot study.

BACKGROUND: Acute otitis media (AOM) is an inflammatory disease of the middle ear causing significant morbidity in early childhood. A pilot study was undertaken to identify the role of various risk factors South Indian children with AOM, especially the role of nasopharyngeal otopathogens. METHODOLOGY: A prospective case control pilot study was conducted in children aged below six years, presenting to a single tertiary care from 2018 to 2019. Fifty cases with AOM and 45 age and gender matched controls were recruited. Two nasopharyngeal swabs were collected, one was processed for bacterial culture. The other swab was processed according to the CDC recommended broth enrichment method to identify carriage of S. pneumoniae. Subsequent serotyping was done by Quellung method and conventional sequential multiplex PCR. RESULT: Otalgia was the major presentation seen in 92% of the children with AOM. None of the clinical and demographic characteristics were found to be statistically significant between the cases and controls. The most common otopathogen was S. pneumoniae (55%) followed by H. influenza (29%). The common S. pneumoniae serotypes encountered were 11A and 19F.Nasopharyngeal colonization with S. pneumoniae [OR 6.57, p < 0.003] and H. influenzae [OR14.18, p < 0.003] were significant risk factors for AOM in children. The risk increased with co-colonization (OR 13.89,p < 0.003). CONCLUSION: This study strengthens the significant association between nasopharyngeal colonization of otopathogens and AOM as a risk factor that is enhanced by co-colonization.S. pneumoniae was the main otopathogen in this population, serotypes 11A and 19F being the most common.

Antimicrobial Photodynamic Therapy with Chlorin e6 Is Bactericidal against Biofilms of the Primary Human Otopathogens.

Moraxella catarrhalis, Streptococcus pneumoniae, and nontypeable Haemophilus influenzae (NTHi) are ubiquitous upper respiratory opportunistic pathogens. Together, these three microbes are the most common causative bacterial agents of pediatric otitis media (OM) and have therefore been characterized as the primary human otopathogens. OM is the most prevalent bacterial infection in children and the primary reason for antibiotic administration in this population. Moreover, biofilm formation has been confirmed as a primary mechanism of chronic and recurrent OM disease. As bacterial biofilms are inherently metabolically recalcitrant to most antibiotics and these complex structures also present a significant challenge to the immune system, there is a clear need to identify novel antimicrobial approaches to treat OM infections. In this study, we evaluated the potential efficacy of antibacterial photodynamic therapy (aPDT) with the photosensitizer chlorin e6 (Ce6) against planktonic as well as biofilm-associated M. catarrhalis, S. pneumoniae, and NTHi. Our data indicate aPDT with Ce6 elicits significant bactericidal activity against both planktonic cultures and established biofilms formed by the three major otopathogens (with an efficacy of ≥99.9% loss of viability). Notably, the implementation of a novel, dual-treatment aPDT protocol resulted in this disinfectant effect on biofilm-associated bacteria and, importantly, inhibited bacterial regrowth 24 h posttreatment. Taken together, these data suggest this novel Ce6-aPDT treatment may be a powerful and innovative therapeutic strategy to effectively treat and eradicate bacterial OM infections and, significantly, prevent the development of recurrent disease.IMPORTANCE Otitis media (OM), or middle ear disease, is the most prevalent bacterial infection in children and the primary reason for antibiotic use and surgical intervention in the pediatric population. Biofilm formation by the major bacterial otopathogens, Moraxella catarrhalis, Streptococcus pneumoniae, and nontypeable Haemophilus influenzae, has been shown to occur within the middle ears of OM patients and is a key factor in the development of recurrent disease, which may result in hearing impairment and developmental delays. Bacterial biofilms are inherently impervious to most antibiotics and present a significant challenge to the immune system. In this study, we demonstrate that antimicrobial photodynamic therapy (aPDT) using the photosensitizer chlorin e6 elicits significant bactericidal activity versus planktonic and biofilm-associated otopathogens and supports further analyses of this novel, efficacious, and promising technology as an adjunctive treatment for acute and recurrent OM.

Niche- and Gender-Dependent Immune Reactions in Relation to the Microbiota Profile in Pediatric Patients with Otitis Media with Effusion.

Otitis media with effusion (OME) is a common inflammatory disease that primarily affects children. OME is defined as a chronic low-grade inflammation of the middle ear (ME), without any signs of infection and with effusion persisting in the ME for more than 3 months. The precise pathogenesis is, however, not fully understood. Here, we comprehensively characterized and compared the host immune responses (inflammatory cells and mediators) and the overall microbial community composition (microbiota) present in matched middle ear effusion (MEE) samples, external ear canal (EEC) lavages, and nasopharynx (NPH) samples from children with OME. Female patients had significantly increased percentages of T lymphocytes and higher levels of a wide array of inflammatory mediators in their MEE compared to that of male patients, which were unrelated to microbiota composition. The relative abundances of identified microorganisms were strongly associated with their niche of origin. Furthermore, specific inflammatory mediators were highly correlated with certain bacterial species. Interestingly, some organisms displayed a niche-driven inflammation pattern in which presence of Haemophilus spp. and Corynebacterium propinquum in MEE was accompanied by proinflammatory mediators, whereas their presence in NPH was accompanied by anti-inflammatory mediators. For Turicella and Alloiococcus, we found exactly the opposite results, i.e., an anti-inflammatory profile when present in MEE, whereas their presence in the the NPH was accompanied by a proinflammatory profile. Together, our results indicate that immune responses in children with OME are highly niche- and microbiota-driven, but gender-based differences were also observed, providing novel insight into potential pathogenic mechanisms behind OME.

Bacterial aetiology of chronic otitis media with effusion in children - risk factors.

BACKGROUND: Otitis media with effusion (OME) may occur spontaneously because of poor Eustachian tube function or as an inflammatory response following AOM. Bacterial involvement in OME has been widely reported, with various available methods to identify pathogens from middle ear effusion, including traditional culture methods and polymerase chain reaction (PCR). OBJECTIVES: The primary goal of this study was to evaluate the bacteriological profile of middle ear effusion in OME. Risk factors of the bacterial OME aetiology were also identified. METHODS: Middle ear effusions (MEF) from 50 children, aged 2-8 years, diagnosed by ENT and undergoing routine tympanostomy tube placement were collected. MEF samples were streaked on standard microbiological media. Next, DNA was isolated from MEF samples and analysed with multiplex PCR for Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis and Alloiococcus otitidis. RESULTS: In multiplex PCR assay 37 (74%) of 50 children were positive for at least one of the four microorganisms. In 27.0% positive children multiple bacterial pathogens were identified. A. otitidis was the most frequently identified in positive MEF children (59.5%). By multiplex PCR, H. influenzae, S. pneumoniae and M. catarrhalis were detected in 24, 18 and 8% of OME patients, respectively. There was significant association between bilateral infection and H. influenzae aetiology of OME. CONCLUSIONS: Overall we found OME predominantly a single otopathogen infection caused mainly by A. otitidis, which is difficult in identification using standard culture method, ahead to S. pneumoniae and H. influenzae. However, one third of MEF samples had multiple bacterial pathogens.

Subversion of host immune responses by otopathogens during otitis media.

Otitis media (OM) is one of the most common ear diseases affecting humans. Children are at greater risk and suffer most frequently from OM, which can cause serious deterioration in the quality of life. OM is generally classified into two main types: acute and chronic OM (AOM and COM). AOM is characterized by tympanic membrane swelling or otorrhea and is accompanied by signs or symptoms of ear infection. In COM, there is a tympanic membrane perforation and purulent discharge. The most common pathogens that cause AOM are Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis whereas Pseudomonas aeruginosa and Staphylococcus aureus are commonly associated with COM. Innate and adaptive immune responses provide protection against OM. However, pathogens employ a wide arsenal of weapons to evade potent immune responses and these mechanisms likely contribute to AOM and COM. Immunologic evasion is multifactorial, and involves damage to host mucociliary tract, genetic polymorphisms within otopathogens, the number and variety of different otopathogens in the nasopharynx as well as the interaction between the host's innate and adaptive immune responses. Otopathogens utilize host mucin production, phase variation, biofilm production, glycans, as well as neutrophil and eosinophilic extracellular traps to induce OM. The objective of this review article is to discuss our current understanding about the mechanisms through which otopathogens escape host immunity to induce OM. A better knowledge about the molecular mechanisms leading to subversion of host immune responses will provide novel clues to develop effective treatment modalities for OM.

Determination of Biofilm-Forming Capacity of Otopathogens Isolated from Discharging Ears in Children with Chronic Otitis Media.

Chronic otitis media is a common disease of the developing world with persistent ear discharge, leading to major complications. This study describes the microorganisms isolated from the middle ear and nasopharynx of children with chronically discharging ears. Middle ear and nasopharyngeal swabs from 89 children were studied, and the microorganisms isolated were assessed for biofilm-forming ability. Methicillin-susceptible Staphylococcus aureus was common in the nasopharynx, while the middle ear showed predominantly pseudomonas and Methicillin-resistant S. aureus. Pseudomonas aeruginosa showed strong biofilm formation, whereas Escherichia coli, Proteus sp. and Providentia sp. were weak biofilm producers. S. aureus isolates were negative for biofilm formation.