Otitis media is an extremely common pediatric ailment caused by opportunists that reside within the nasopharynx. Inflammation within the upper airway can promote ascension of these opportunists into the middle ear chamber. Otitis media can be chronic/recurrent in nature, and a wealth of data indicates that in these cases the bacteria persist within biofilms. Epidemiological data demonstrates most cases of otitis media are polymicrobial, which may have significant impact on antibiotic resistance. In this study, we used in vitro biofilm assays and rodent infection models to examine the impact of polymicrobial infection with Moraxella catarrhalis and Streptococcus pneumoniae (pneumococcus) on biofilm resistance to antibiotic treatment and persistence in vivo. Consistent with prior work, M. catarrhalis conferred beta-lactamase dependent passive protection from beta-lactam killing to pneumococci within polymicrobial biofilms. Moreover, pneumococci increased resistance of M. catarrhalis to macrolide killing in polymicrobial biofilms. However, pneumococci increased colonization in vivo by M. catarrhalis in a quorum signal-dependent manner. We also found that co-infection with M. catarrhalis affects middle ear ascension of pneumococci in both mice and chinchillas. Therefore, we conclude that residence of M. catarrhalis and pneumococci within the same biofilm community significantly impacts resistance to antibiotic treatment and bacterial persistence in vivo.

Even in the vaccine era, Streptococcus pneumoniae (the pneumococcus) remains a leading cause of otitis media, a significant public health burden, in large part because of the high prevalence of nasal colonization with the pneumococcus in children. The primary pneumococcal neuraminidase, NanA, which is a sialidase that catalyzes the cleavage of terminal sialic acids from host glycoconjugates, is involved in both of these processes. Coinfection with influenza A virus, which also expresses a neuraminidase, exacerbates nasal colonization and disease by S. pneumoniae, in part via the synergistic contributions of the viral neuraminidase. The specific role of its pneumococcal counterpart, NanA, in this interaction, however, is less well understood. We demonstrate in a mouse model that NanA-deficient pneumococci are impaired in their ability to cause both nasal colonization and middle ear infection. Coinfection with neuraminidase-expressing influenza virus and S. pneumoniae potentiates both colonization and infection but not to wild-type levels, suggesting an intrinsic role of NanA. Using in vitro models, we show that while NanA contributes to both epithelial adherence and biofilm viability, its effect on the latter is actually independent of its sialidase activity. These data indicate that NanA contributes both enzymatically and nonenzymatically to pneumococcal pathogenesis and, as such, suggest that it is not a redundant bystander during coinfection with influenza A virus. Rather, its expression is required for the full synergism between these two pathogens.

Streptococcus pneumoniae (pneumococcus) is both a widespread nasal colonizer and a leading cause of otitis media, one of the most common diseases of childhood. Pneumococcal phase variation influences both colonization and disease and thus has been linked to the bacteria's transition from colonizer to otopathogen. Further contributing to this transition, coinfection with influenza A virus has been strongly associated epidemiologically with the dissemination of pneumococci from the nasopharynx to the middle ear. Using a mouse infection model, we demonstrated that coinfection with influenza virus and pneumococci enhanced both colonization and inflammatory responses within the nasopharynx and middle ear chamber. Coinfection studies were also performed using pneumococcal populations enriched for opaque or transparent phase variants. As shown previously, opaque variants were less able to colonize the nasopharynx. In vitro, this phase also demonstrated diminished biofilm viability and epithelial adherence. However, coinfection with influenza virus ameliorated this colonization defect in vivo. Further, viral coinfection ultimately induced a similar magnitude of middle ear infection by both phase variants. These data indicate that despite inherent differences in colonization, the influenza A virus exacerbation of experimental middle ear infection is independent of the pneumococcal phase. These findings provide new insights into the synergistic link between pneumococcus and influenza virus in the context of otitis media.

Two research questions are explored in this paper: what are the specific leader activities that contribute most to successful change in the Royal Air Force? And, what are the specific dimensions of leadership that contribute to successful change in the Royal Air Force? The results of a study into the part that leadership plays in successful change in the Royal Air Force (RAF) are presented. The Leadership Dimensions questionnaire was used to measure the dimensions of leadership and change contexts, and a 3608 feedback questionnaire gathered data on leader activities and the success of change programmes. The sample population was senior management in the RAF with 3608 feedback from colleagues. Hierarchies of the leader activities and dimensions of leadership are presented to identify the components that exert the strongest influence on the success of leading change in the RAF. Managing resources, engaging communication and empowering were identified as the three dimensions that contribute most towards successful change; and 'creating a clear vision', 'adjusting the work culture to meet the long term needs of the change' and 'generating early successes' were the three leader activities that exhibited the strongest influence on success. Recommendations are presented for future research in this area and for the RAF, based on these findings.

Infection with influenza A virus (IAV) can lead to increased susceptibility to subsequent bacterial infection, often with Streptococcus pneumoniae. Given the substantial modification of the lung environment that occurs following pathogen infection, there is significant potential for modulation of immune responses. Here we show that coinfection of mice with influenza virus followed by the non-invasive EF3030 strain of Streptococcus pneumoniae leads to a significant decrease in the virus specific CD8+ T cell response in the lung. Adoptive transfer studies suggest this reduction contributes to disease in coinfected animals. The reduced number of lung effector cells in coinfected animals was associated with increased death as well as a reduction in cytokine production in surviving cells. Further, cells that retained the ability to produce interferon γ exhibited a decreased potential for co-production of tumor necrosis factor α. Reduced cytokine production was directly correlated with a decrease in the level of mRNA. Negative regulation of cells in the mediastinal lymph node (MLN) was minimal compared to that present in the lung, supporting a model of negative regulation in the tissue harboring high pathogen burden. These results elucidate a new aspect of immune regulation as a result of entry of a coinfecting pathogen, modulation of ongoing adaptive immune responses in the lung. These findings reveal a novel dynamic interplay between concurrently infecting pathogens and the adaptive immune system.

Objective: To determine the suitability of four research methods to measure the rate of child restraint device (CRD) use and incorrect use in New Zealand and obtain data on barriers to CRD use. Design and setting: To assess the rates of CRD use among vehicles carrying children 8 years of age and under, two methods were piloted-namely, an unobtrusive observational survey and a short interview and close inspection. A self administered questionnaire and focus group interviews were also piloted to assess CRD use, reasons for use and non-use, and to obtain information on barriers to their use. Respondents to all methods except the focus groups were approached in supermarket car park sites at randomly selected times. Focus groups were established with parents identified through early childhood organisations. All methods were assessed on criteria related to efficiency, representativeness, and ability to obtain the necessary data. Results: The observational survey provided a simple method for identifying rates of CRD use, while the self administered questionnaire obtained data on demographic characteristics and reported the installation and use/non-use of CRDs. The interview/inspection addressed all the questions of both the above methods and enabled incorrect CRD use to be examined. The focus groups provided the most meaningful information of all methods on barriers to CRD use. Discussion and conclusion: Advantages and limitations of these methods are discussed and some refinements of the original instruments are proposed. The interview/inspection and focus group methods were identified as being more appropriate for efficiently obtaining reliable data on CRD use and identification of barriers to CRD use.

Adenoviral infection is a major risk factor for otitis media. We hypothesized that adenovirus promotes bacterial ascension into the middle ear through the disruption of normal function in the Eustachian tubes due to inflammation-induced changes. An intranasal infection model of the chinchilla was used to test the ability of type 5 adenovirus to promote middle ear infection by Streptococcus pneumoniae. The hyper-inflammatory adenovirus mutant dl327 and the non-replicating adenovirus mutant H5wt300ΔpTP were used to test the role of inflammation and viral replication, respectively, in promotion of pneumococcal middle ear infection. Precedent infection with adenovirus resulted in a significantly greater incidence of middle ear disease by Streptococcus pneumoniae as compared to non-adenovirus infected animals. Infection with the adenovirus mutant dl327 induced a comparable degree of bacterial ascension into the middle ear as did infection with the wild-type virus. By contrast, infection with the non-replicating adenovirus mutant H5wt300ΔpTP resulted in less extensive middle ear infection compared to the wild-type adenovirus. We conclude that viral replication is necessary for adenoviral-induced pneumococcal middle ear disease.

Streptococcus pneumoniae strains lacking capsular polysaccharide have been increasingly reported in carriage and disease contexts. Since most cases of otitis media involve more than one bacterial species, we aimed to determine the capacity of a nonencapsulated S. pneumoniae clinical isolate to induce disease in the context of a single-species infection and as a polymicrobial infection with nontypeable Haemophilus influenzae. Using the chinchilla model of otitis media, we found that nonencapsulated S. pneumoniae colonizes the nasopharynx following intranasal inoculation, but does not readily ascend into the middle ear. However, when we inoculated nonencapsulated S. pneumoniae directly into the middle ear, the bacteria persisted for two weeks post-inoculation and induced symptoms consistent with chronic otitis media. During coinfection with nontypeable H. influenzae, both species persisted for one week and induced polymicrobial otitis media. We also observed that nontypeable H. influenzae conferred passive protection from killing by amoxicillin upon S. pneumoniae from within polymicrobial biofilms in vitro. Therefore, based on these results, we conclude that nonencapsulated pneumococci are a potential causative agent of chronic/recurrent otitis media, and can also cause mutualistic infection with other opportunists, which could complicate treatment outcomes.