Moraxella catarrhalis, Kingella , and Other Gram-Negative Cocci

History

M. catarrhalis has an interesting and checkered taxonomic history. The bacterium was first described a century ago and was suspected by Sir William Osler to be the cause of his own terminal pneumonia. After having been initially named Micrococcus catarrhalis , the organism's name was subsequently changed to Neisseria catarrhalis because of its similarities in phenotype and ecologic niche to Neisseria spp. In 1970 it was transferred to the new genus Branhamella on the basis of differences in fatty-acid content and DNA hybridization studies, compared with other Neisseriaceae. The name Moraxella catarrhalis was subsequently proposed, and this is the most widely accepted name at this time.

For most of the last century M. catarrhalis was regarded as an upper respiratory tract commensal. However, since the late 1970s, investigators from many centers have accumulated compelling evidence that M. catarrhalis is an important and common respiratory tract pathogen in humans.

Microbiology

Current taxonomic classification schemes include three genera in the family Moraxellaceae —Moraxella, Acinetobacter , and Psychrobacter. M. catarrhalis is a gram-negative diplococcus that is indistinguishable from Neisseria by Gram stain. The organism grows well on blood agar, chocolate agar, and a variety of media. M. catarrhalis are difficult to distinguish from Neisseria by colony morphology, particularly after overnight growth on agar plates. After 48 hours of growth, M. catarrhalis colonies tend to be larger than Neisseria and take on a pink color. In addition, colonies display the hockey puck sign by sliding along the surface of the agar when pushed. Because samples from the respiratory tract frequently contain Neisseria , suspicious colonies should be tested for the possibility that they are M. catarrhalis. The similarity in colony morphology between commensal Neisseria and M. catarrhalis results in the underestimation of M. catarrhalis in cultures of human respiratory tract samples. M. catarrhalis produces oxidase, catalase, and DNase. Several kits to speciate M. catarrhalis are commercially available.

Epidemiology and Respiratory Tract Colonization

M. catarrhalis has been recovered exclusively from humans. The prevalence of colonization is highly dependent on age. The upper respiratory tract of approximately 1% to 5% of healthy adults is colonized by M. catarrhalis. By contrast, nasopharyngeal colonization with M. catarrhalis is common throughout infancy. Some published studies have shown a higher rate of colonization during winter months; this higher rate may be a result of the appearance of respiratory viral illnesses during colder months. Substantial regional differences in colonization rates are observed. For example, 66% of infants in a study in Buffalo, New York, were colonized during the first year of life, whereas a similar study in Göteborg, Sweden, showed a colonization rate of approximately half of that level. A study of rural Aboriginal infants near Darwin, Australia revealed that 100% of infants were colonized by M. catarrhalis by the age of 3 months. The explanation for the marked differences in rates of colonization is not yet known. Several factors, including living conditions, daycare, crowding, hygiene, environmental factors (e.g., household smoking), genetic characteristics of the populations, and host factors, may play a role. Microbial interactions in the nasopharynx among cocolonizing respiratory pathogens, including viruses and bacteria, and normal flora contribute to the complexity and dynamics of nasopharyngeal colonization. The widespread use of pneumococcal conjugate vaccines has caused changes in patterns of nasopharyngeal colonization by reducing colonization with vaccine serotypes of Streptococcus pneumoniae. This effect has resulted in “replacement” of vaccine serotypes of S. pneumoniae by nonvaccine pneumococcal serotypes. Whether replacement by nontypeable Haemophilus influenzae and M. catarrhalis also occurs will require careful monitoring.

Nasopharyngeal colonization with middle ear pathogens, including M. catarrhalis , is associated with otitis media. Early colonization is a risk factor for recurrent otitis media. Otitis-prone children are colonized with M. catarrhalis at a higher rate compared with healthy children.

M. catarrhalis is isolated from the sputum of adults with chronic obstructive pulmonary disease (COPD). Approximately 50% of episodes of acquisition of M. catarrhalis are associated with acute exacerbations of COPD. Adults with COPD develop systemic and mucosal immune responses and clear M. catarrhalis from the respiratory tract efficiently after a relatively short duration (<1 month). A smaller proportion of episodes of colonization may persist for up to 2 years. Acquisition and clearance of the organism results in strain-specific protection.

Pathogenesis

M. catarrhalis causes mucosal infections in children and adults. The pathogenesis of infection appears to involve contiguous spread of the bacterium from its colonizing position in the respiratory tract to cause clinical signs of infection. In the case of otitis media the isolates recovered from the middle ear are present in the nasopharynx, indicating that the middle ear isolate came from the nasopharynx via the eustachian tube. Colonization of the upper respiratory tract with middle ear pathogens, including M. catarrhalis , is a necessary first step in the pathogenesis of otitis media. However, colonization alone is not sufficient to cause disease. An inciting event, such as a viral infection, in a child colonized with a middle ear pathogen is probably necessary for bacteria to move to the middle ear and cause otitis media. In the case of infection in adults with COPD the acquisition of a new strain is critical in the pathogenesis of infection.

A key step in the initiation of infection is adherence of M. catarrhalis to the human respiratory epithelium. Several adhesins with varying specificities for host cells have been identified ( Table 213.1 ). Some of these surface antigens and others are being evaluated as potential vaccine antigens. In addition to adherence to the epithelial surface, M. catarrhalis also resides within and beneath the epithelium and invades host cells. Indeed, colonization of the nasopharynx by M. catarrhalis is more frequent than is revealed by surface culture.

The outer membrane of M. catarrhalis contains lipooligosaccharide (LOS). LOS consists of a lipid A core coupled to oligosaccharides. The structure of the LOS resembles that of other nonenteric gram-negative bacteria in that the molecule lacks the long polysaccharide side chains observed in enteric gram-negative bacteria. Three major antigenic types of LOS can be distinguished, accounting for 95% of all strains. The different serotypes are based on differences in terminal sugars in the LOS molecule. A detoxified LOS molecule is a potential vaccine antigen.

The interaction of selected surface antigens with receptors of various host cells in the respiratory tract has important effects in mediating host responses to M. catarrhalis. For example M. catarrhalis is mitogenic for B lymphocytes through interaction with the MID/Hag protein. The surface protein UspA2 regulates nuclear factor kappa B and subsequent interleukin-8 release by human respiratory epithelial cells. LOS stimulates human monocytes to produce proinflammatory cytokines in both Toll-like receptor 4– and CD14-dependent pathways. Like many gram-negative bacteria, M. catarrhalis sheds vesicles from its surface; these vesicles are internalized by respiratory epithelial cells and mediate several virulence mechanisms, including B-cell activation, induction of inflammation, and delivery of β-lactamase, which may promote survival of copathogens. The availability of the genome sequence of clinical isolates is facilitating studies of pathogenesis. M. catarrhalis evades host immunity through specific binding of several human extracellular matrix molecules in connective tissue, including vitronectin, fibronectin, collagen, plasminogen, and cartilage oligomeric matrix protein. These host molecular patterns become exposed when epithelial integrity is interrupted, for example, by chronic inflammation or viral infection.

A reliable animal model that parallels human infection has not yet been developed for M. catarrhalis. The specificity of M. catarrhalis for humans creates challenges for the development of a useful model to study pathogenesis. The chinchilla model of otitis media is used widely to study otitis media caused by other bacteria, but chinchillas readily clear M. catarrhalis when it is instilled into the middle ear. M. catarrhalis colonizes the nasopharynx of the chinchilla, so this model may prove useful. The most widely used model is a mouse pulmonary clearance model that measures the rate of clearance of M. catarrhalis from the lungs after intratracheal challenge. This model does not parallel human infection but has been used as a guide to identify and study potential vaccine antigens. Models that involve coinfection of M. catarrhalis with other viral and respiratory tract pathogens may prove to be useful in understanding the role of M. catarrhalis as a copathogen.

Clinical Manifestations

Otitis Media

Approximately 80% of children experience at least one episode of acute otitis media by the age of 3 years. A subset of children experiences recurrent otitis media, which is associated with a delay in speech and language development. Careful studies from many centers have defined the cause of acute otitis media by culturing middle ear fluid obtained by tympanocentesis, which has been considered the gold standard for determining the etiology of otitis media. Although some differences among studies are observed, the results from centers in the United States and Europe are remarkably consistent in showing that S. pneumoniae , nontypeable H. influenzae , and M. catarrhalis are the predominant bacterial causes of acute otitis media. Overall, based on cultures of middle ear fluid, approximately 5% to 20% of cases of acute otitis media are caused by M. catarrhalis . However, culture detects pathogens in only a subset of otitis media. Using more sensitive molecular analysis, M. catarrhalis is detected alone or with other pathogens in 30% to 50% of middle ear fluid samples from children with otitis media ( Fig. 213.1 ). Analysis of middle ear fluid by PCR increases the frequency of detection of S. pneumoniae and nontypeable H. influenzae 3.2-fold compared with culture, whereas M. catarrhalis is 4.5 times more likely to be identified by PCR. Several lines of evidence indicate that the presence of bacterial DNA in clinical samples indicates active infection, including (1) detection of messenger RNA, (2) viable bacteria in biofilms, and (3) studies in the chinchilla model. When M. catarrhalis is isolated from middle ear fluid in acute otitis media, it is present as a sole pathogen in approximately 75% of cases and as a copathogen in the remaining 25%. The widespread use of pneumococcal conjugate vaccines is altering the distribution of pathogens in otitis media, emphasizing the importance of continuous monitoring.

Lower Respiratory Tract Infections in Chronic Obstructive Pulmonary Disease

M. catarrhalis causes lower respiratory tract infections in adults, particularly in the setting of COPD. The recognition of M. catarrhalis as a pathogen in this setting was delayed until the past 20 years because M. catarrhalis is indistinguishable from commensal Neisseria by Gram stain and difficult to distinguish by colony morphology. Therefore, unless clinical microbiology laboratories specifically test colonies that appear to be Neisseria, M. catarrhalis will be missed as a potential pathogen in sputum.

Several lines of evidence have established that M. catarrhalis causes exacerbations of COPD :

• Analysis of sputum samples of a subset of patients with exacerbations of COPD demonstrates a predominance of gram-negative diplococci on Gram stain and almost pure cultures of M. catarrhalis.

• Studies using transtracheal aspiration and bronchoscopy with the protected specimen brush to sample the lower airways have revealed pure cultures of M. catarrhalis in some patients with exacerbations of COPD.

• The levels of inflammatory markers in the sputum of patients with exacerbations with positive cultures for M. catarrhalis are higher than the level in sputum of culture-negative exacerbations.

• A specific immune response has been observed after exacerbations of COPD associated with M. catarrhalis in the sputum.

• Acquisition of a new strain of M. catarrhalis is associated with clinical exacerbation.

• Exacerbations resulting from acquisition of a new strain of M. catarrhalis are associated with protease-antiprotease imbalance in the airways.

M. catarrhalis causes approximately 10% of exacerbations of COPD, making the bacterium the second most common bacterial cause of exacerbations after nontypeable H. influenzae .

The clinical manifestations of exacerbations of COPD caused by M. catarrhalis are similar to those of exacerbations caused by other bacteria, such as nontypeable H. influenzae. Patients experience increased cough and sputum production, increased sputum purulence, and increased dyspnea compared with baseline symptoms. Sputum Gram staining shows intracellular and extracellular gram-negative diplococci as the exclusive or predominant bacterial form ( Fig. 213.2 ), and cultures grow predominantly M. catarrhalis.