Attachment of Nontypable Haemophilus influenzae to Human Pharyngeal Epithelial Cells Mediated by a Ganglioside Receptor

Nontypable Haemophilus influenzae (NTHI) is one of the major pathogens of human respiratory infections and has the ability to attach to pharyngeal epithelial cells. We characterized the epithelial cell receptor to which NTHI bind. Neuraminidase pretreatment of pharyngeal epithelial cells resulted in a significant decrease in NTHI attachment, suggesting sialic acid as an important component of the receptor. The attachment was not decreased in NTHI pretreated with 1,000 μg/ml of fucose, N-acetyl-neuraminic acid, N-acetyl-glucosamine, N-acetyl-galactosamine, acetyl-salicylic acid and colominic acid. Only treatment with gangliosides D1a, D1b and D2 at a concentration of 12.5 μg/ml significantly decreased the attachment. On the other hand, treatment with gangliosides M1, M2, M3, D3, T1b and asialoganglioside M1 did not decrease the attachment of NTHI. Only ganglioside D2 inhibited the attachment significantly at a concentration of 12.5 ng/ml. Other isolates of NTHI showed a decrease in attachment after treatment with ganglioside D2. Treatment of cells with anti-human GD2 monoclonal antibody also decreased the attachment of NTHI in a dose-dependent manner. This study indicates that sialic acid glycoconjugate, GD2, is one of the receptors of NTHI on human pharyngeal epithelial cells.     

Prevalence of hicAB, lav, traA, and hifBC among Haemophilus influenzae middle ear and throat strains

Nontypeable Haemophilus influenzae (NTHi) is an important cause of illness among children. To further understand the role of laterally transferred genes in NTHi colonization and otitis media, the prevalence of hicAB, lav, tnaA, and hifBC was determined among 44 middle ear and 35 throat NTHi isolates by dot-blot hybridization.     

Neutrophil Response to Nontypable Haemophilus influenzae in Respiratory Infections

Sputa from patients with respiratory infections by nontypable Haemophilus influenzae (H. influenzae) were investigated by electron microscopy. The cell wall of H. influenzae appeared wavy and nonwavy. In the cell wall the peptidoglycan layer was ill-defined. These patients had adequate IgG response in the serum against H. influenzae. However neither capsule nor fimbriae were found. Different stages of phagocytosis and destruction of the bacteria by polymorphonuclear neutrophils (PMN) were observed. PMNs were also found to phagocytose the debris. Evidences were found that the debris is formed mainly by the destruction of polymorphonuclear neutrophil. Extracellular lysosomes were also observed, which may have a role in destruction of both bacteria and host tissue. It was concluded that nontypable H. influenzae are nonfimbriated and noncapsulated during infection. Debris are the end product of PMN destruction, and phagocytosis of debris by PMNs has a role in the pathogenesis of chronic respiratory diseases.     

Role of Toll-like receptor 4 in innate immune responses in a mouse model of acute otitis media

Nontypeable Haemophilus influenzae (NTHi) is considered a major pathogen underlying middle ear infection. This study characterized the role of Toll-like receptor 4 in the innate immune responses to acute otitis media induced by NTHi in mice. We used C3H/HeJ mice, which have nonfunctional Toll-like receptor 4, and normal wild-type C3H/HeN mice. NTHi were injected into the tympanic bulla, and middle ear effusions and tissues were collected. In C3H/HeN mice, the severity of acute otitis media decreased promptly with a significant reduction in bacterial recovery from middle ear effusions 48 h after injection. In contrast, all C3H/HeJ mice had otitis media at all time points examined, and increasing bacterial counts from middle ear effusions were detected in C3H/HeJ mice 72 h after injection. Expression of intracellular adhesion molecule-1 by the middle ear mucosa paralleled the number of polymorphonuclear cells in the middle ear in both strains. The findings of transmission electron microscopy revealed that phagocytosis and phagosome maturation of polymorphonuclear cells was impaired in C3H/HeJ mice. Our findings indicate that Toll-like receptor 4 plays a part in the early accumulation and functional promotion of polymorphonuclear cells in the middle ear for eradicating NTHi infection.     

Antibodies directed at a conserved motif in loop 6 of outer membrane protein P2 of nontypeable Haemophilus influenzae recognize multiple strains in immunoassays

The P2 porin is the most abundant protein in the outer membrane of nontypeable Haemophilus influenzae. Analysis of P2 sequences from a limited number of strains reveals the presence of both heterogeneous and conserved surface-exposed loops of the P2 molecule among strains. We have previously shown that antibodies raised against the loop 6 sequence of P2 from strain 5657 are bactericidal against multiple isolates. In this study, we determined the nucleotide sequence of the loop 6 region of the P2 molecule from 108 strains of nontypeable H. influenzae in order to assess more rigorously the degree of conservation of loop 6. Based on this analysis, we identified a conserved sequence, different from that of strain 5657, that occurs in approximately one-third of the strains sequenced. To assess the potential of this peptide as a vaccine antigen, antibodies raised to a multiple antigenic peptide corresponding to this sequence were characterized with respect to specificity for the P2 molecule and reactivity with heterologous strains in immunoblot assay, flow cytometry and bactericidal assays. Antibodies were reactive to the P2 molecule of 16 of 20 strains tested by immunoblot assay. Antibodies recognized nine of the 20 strains in a flow cytometry assay, and 13 of 20 demonstrated complement-mediated killing in bactericidal assays. These results support the concept of using conserved regions of the P2 protein as a vaccine antigen.     

Development of antibodies against the putative proteinase maturation protein A in relation to pneumococcal carriage and otitis media

The putative pneumococcal proteinase maturation protein A is a potential pneumococcal vaccine candidate. We examined serum antipneumococcal proteinase maturation protein A antibodies at 6, 12, 18 and 24 months of age, and showed that the age-related development of antipneumococcal proteinase maturation protein A antibodies is associated with pneumococcal contacts. A higher antipneumococcal proteinase maturation protein A antibody concentration at 18 months of age tends to predict for a lower risk of pneumococcal acute otitis media in the following 6 months (relative risk: 0.84, 95% confidence interval: 0.62-1.13).     

Isolation of Alloiococcus otitidis from Indigenous and non-Indigenous Australian children with chronic otitis media with effusion

During the last decade Alloiococcus otitidis has been identified in specimens from patients with chronic otitis media with effusion. Whereas most of those studies employed molecular techniques, we used minor modifications of conventional microbiological methods to isolate and identify A. otitidis in samples obtained from 20/50 (40%) children referred for myringotomy. Alloiococcus otitidis was isolated from 10/22 (45%) Indigenous and 10/28 (36%) non-Indigenous children. This is the first report of isolation of A. otitidis from Australian children with chronic otitis media. All isolates were sensitive to penicillin, but 14/20 (70%) of the isolates were resistant or partially resistant to erythromycin as assessed by the E-test.     

Evidence of local antibody response against Alloiococcus otitidis in the middle ear cavity of children with otitis media

Alloiococcus otitidis is a recently discovered bacterium frequently associated with otitis media. However, no study is available as to whether A. otitidis has a pathogenic role and induces local immune response in the middle ear as a true pathogen. Whole bacterial sonicate of A. otitidis was separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and transferred to a nitrocellulose membrane. Then, Western blot analysis was performed with supernatant of the middle ear effusions from children with A. otitidis-positive otitis media. SDS-PAGE of the bacterial sonicate showed several protein bands, designated A1-A11. Western blot analysis revealed the presence of IgG, secretory IgA, IgG2, and IgM against A. otitidis in the middle ear effusions. Absorption of the specimens with sonicates of other major middle ear pathogens did not alter the reactivity of antibodies against the alloiococcal antigens. The results suggest that specific local immune response against A. otitidis is induced during middle ear infection of the organism as a true pathogen. A5, A6 or A11 is expected to be a main antigenic determinant. This is the first report to show evidence of local antibody response against A. otitidis and to disclose antigenic components of A. otitidis.     

儿童腺样体肥大程度与分泌性中耳炎发生的相关性研究(英文)

目的:探讨儿童腺样体肥大程度与分泌性中耳炎发生及预后的相关性,指导临床医师对分泌性中耳炎作出早期诊断和治疗。方法:239例住院手术切除腺样体的儿童,常规行鼻咽侧位片、声导抗检查;部分伴耳部症状、声导抗显示C型曲线或查体可疑鼓室积液征者行颞骨CT检查或术中行鼓室穿刺。经统计学分析,比较分泌性中耳炎与腺样体肥大程度及咽鼓管咽口情况的相关性。结果:在239例腺样体肥大儿童中,经鼓室穿刺证实合并分泌性中耳炎者34例(63耳,14.2%),其中鼓室曲线呈B型者33耳(52.4%),C型(-200 dapa)者10耳(15.9%),C型(-200 dapa)者20耳(31.7%)。结果表明分泌性中耳炎的发生与腺样体肥大程度及咽鼓管园枕受压迫的程度呈正相关。结论:声导抗检查不能作为分泌性中耳炎诊断的金标准,必要时可行颞骨CT明确诊断;对腺样体肥大伴分泌性中耳炎的儿童鼻内镜下腺样体切除为其主要疗法,配合鼓室穿刺多可治愈,对反复发作的分泌性中耳炎行鼓室置管术,避免术后并发症的发生。     

Mastoiditis--paleopathological evidence of a rarely reported disease

Since antibiotics have become available, mastoiditis has become a rare disease in modern Western societies. However, it is still common in developing countries. It can be hypothesized that in earlier historical and prehistoric times, mastoiditis must have posed a serious threat to people's lives, and that the prevalence of this disease is probably underrepresented in the paleopathological literature. The present study identifies pathological changes in the pneumatized cells of the mastoid process in human skeletal samples from two early medieval cemeteries from Germany (Dirmstein: n = 152 mastoids, Rhens: n = 71 mastoids), using macroscopic, endoscopic, low-power microscopic, scanning-electron and light microscopic techniques, and draws some epidemiological conclusions as to the frequency of the disease diagnosed in the archaeological samples. Osseous changes because of mastoiditis were diagnosed in 83.4% of the temporal bones. The frequency in the skeletal sample from Dirmstein was higher than in the sample from Rhens. In both populations, males were more often affected than females and older individuals more often than younger individuals. The high frequency of mastoiditis observed was most likely due to an accumulation of osseous changes during individual lifetimes and supports the hypothesis that mastoiditis was a serious health problem in pre-antibiotic times. It may be assumed that subclinical forms of mastoiditis and their osseous manifestations may even nowadays occur more often than was previously thought. It is suggested that the disease should be given more consideration in paleopathological investigations.     

综合康复治疗对分泌性中耳炎致小儿听力损伤的临床效果

目的:探讨综合康复治疗对分泌性中耳炎致小儿听力损伤的临床效果。方法:以88例2016年1月-2017年8月于我院诊治的分泌性中耳炎致听力损伤患儿为研究对象,将其随机分为综合组和对照组,每组44例。综合组采用综合康复治疗,对照组采用耳部按摩治疗,观察并比较两组临床疗效,治疗前后气导、骨导听阈值的变化。结果:治疗后,综合组有效率为79.55%,显著高于对照组(56.82%,P0.05)。与治疗前相比,两组治疗后0.25-0.8 Hz各频率下气导听阈值及2.0和4.0 kHz下骨导听阈值均显著降低(P0.05),且综合组患儿以上指标均显著低于对照组(P0.05)。结论:综合康复治疗即联合感音训练、耳部按摩、音乐感知及运动训练对分泌性中耳炎致小儿听力损伤有较好的疗效,能改善患儿气导、骨导听阈水平。