Sandbox hemophillus

Pathophysiology

Transmission

• Transmission is by direct contact or by inhalation of respiratory tract droplets.
• Neonates can acquire infection by aspiration of amniotic fluid or contact with genital tract secretions containing the bacteria.

Incubation

Incubation period of Hemophilus influenza infection is variable.

Seeding

• The colonizing bacteria invade the mucosa and enter the bloodstream.
• Eustachian tube dysfunction, antecedent viral upper respiratory tract infection (URTI), foreign bodies, and mucosal irritants, including smoking, can promote infection.
• In patients with underlying chronic obstructive pulmonary disease (COPD) or cystic fibrosis (CF), NTHi frequently colonizes the lower respiratory tract and can exacerbate the disease.

Spread of bacterial load	Type of infection
Eustachian tube	Otitis media
Sinuses	Sinusitis
Respiratory tract	Bronchitis and pneumonia

Pathogenesis

• The antiphagocytic nature of the Hib capsule and the absence of the anticapsular antibody lead to increasing bacterial proliferation.
• When the bacterial concentration exceeds a critical level, it can disseminate to various sites, including meninges, subcutaneous tissue, joints, pleura, pericardia, and lungs.
• The antibody to the Hib capsule plays the primary role in conferring immunity.
• Newborns have a low risk of infection, likely because of acquired maternal antibodies.
• When these transplacental antibodies to the PRP antigen wane, infants are at high risk of developing invasive H influenzae disease, and their immune responses are low even after the disease.
• Therefore, they are at high risk of repeat infections since prior episodes of H influenzae do not confer immunity. By age 5 years, most children have naturally acquired antibodies.
• The Hib conjugate vaccine induces protection by inducing antibodies against the PRP capsule.
• The Hib conjugate vaccine does not provide protection against NTHi strains. Since the widespread use of the Hib conjugate vaccine, NTHi has become more of a pathogen.

Immunological response

Haemophilus influenzae was first identified by Pfeiffer in 1892, who (incorrectly) believed it was the cause of influenza [1]. It is an exclusively human pathogen and was the first bacterium to have its genome completely sequenced. This served as a precursor to the sequencing of the human genome.

H. influenzae is a component of the normal upper respiratory tract flora and is well recognized to be an important cause of systemic infection. It is also a major cause of a variety of respiratory conditions and has had a relatively low profile in this respect in comparison to some other pathogens; such as Mycobacterium tuberculosisand Streptococcuspneumoniae.

Recently there has been increasing recognition that this bacterium has a role in chronic lower respiratory tract inflammation. However the interaction between H. influenzae and the lung is still not well defined. A combination of bacterial pathogenic features and deficiency of host defense may permit this bacterium to establish infection in the lower respiratory tract resulting in inflammation and clinical disease. This revi Mucociliary interactions

The mucociliary apparatus is a first-line structural defense against bacterial infection and H. influenzae strains have a variety of mechanisms which can influence its function. Outer membrane proteins such as P2 and P5 facilitate binding of the bacteria to mucus [11,12]. Lipooligosaccharide (a lipopolysaccharide that lacks the O-side chains and is abbreviated as LOS) is present in the cell wall of NTHi strains and has a significant effect on cilial function; Denny described that LOS produced inhibition of ciliary function and loss/death of ciliary mucosal cells [13]. Similar effects on ciliary function have been described from protein D which is a lipoprotein expressed on the surface of H. influenza[14]. A key step in pathogenesis is the ability to adhere to the respiratory mucosa. NTHi appears to have a preference for nonciliated cells or damaged mucosa. There are several specific mechanisms which NTHi strains use to adhere to mucosa.

Adhesins The adhesins are a common and important factor to facilitate epithelial attachment. These are present on a large proportion of NTHi strains and share some homology with adhesins expressed by Bordatella pertussis[15]. They are also a major target for serum antibodies to NTHi infection [16]. A number of investigators have demonstrated the importance of adhesins and there are 2 main subtypes HMW1 and HMW2 [17].

Pili NTHi express pili which are rod-like projections on the surface which cause agglutination of red blood cells and attachment to respiratory tract epithelial cells [18]. There are 5 different types and these pili are present only on a small subset of strains of NTHi [19-21].

Other factors Approximately 25 % of NTHi strains lack adhesins/pili but are still able to attach efficiently to respiratory epthilium. Two other factors that are important are the Hia and Hap proteins [22].

Evasion of mucosal immunity

After attachment to the mucosal surface, strains of NTHi have a variety of different mechanisms which enhance persistence at the epithelial surface.

Proteases Immunoglobulin (Ig) A is the main antibody subclass that prevents epithelial infection. IgA binds to bacteria and prevents mucosal attachment, inactivates toxins and facilitates cytotoxicity. The predominant IgA subclass is IgA1. NTHi secretes endopeptidases (Types 1 and 2), which cleave and neutralize IgA1 [23,24]. Nearly all NTHi strains express one of these IgA proteases which are highly effective in inhibiting IgA.

Microcolony formation St Geme et al have demonstrated the ability of NTHi to form microcolonies on mucosal surfaces [8]. This property is likely to inhibit the function of secreted bacteriostatic products such as lactoferrrin and lysozymes and also potentially antibody function.

Phase variation/antigenic drift Viruses such as influenza have well-documented abilities to frequently change cell structures and this is a key feature in their pathogenesis. H. influenzae is also able to lose or gain cell structures; a property called phase variation. Structures in which this occurs include LOS, adhesins and pili. This ability has significant implications for the function of antibodies and C-reactive protein [25].

In addition to phase variation, some strains of H. influenzae undergo antigenic drift which involves permanent change in amino acid sequences in some important immune structures/epitopes. This is best described in the context of the outer membrane protein P2 (which is strongly immunogenic) [26]. Other examples of antigenic drift involve P5 outer membrane protein and IgA1 proteases.

Intracellular survival/invasion of local tissue

A potentially very important pathogenic feature of Haemophilus influenzae is its ability to invade local tissue and survive intracellularly in the respiratory tract. This has been described in the context of nontypeable strains. The main cells targeted by NTHi appear to be macrophages and epithelial cells. There have been a number of studies which have demonstrated the in-vitro ability of NTHi to survive inside these cells for at least 72 hours [27-30]. An example of NTHi inside macrophages is shown in Figure 2. o early studies demonstrated the presence of NTHi between the epithelial cells of patients with chronic bronchitis and adenoidal inflammation [30,31]. Electron microscopy showed the disruption of intracellular junctions and the presence of bacteria between cells and in the phagocytic vacuoles of mononuclear cells. Forsgren et al demonstrated the presence of viable NTHi in mononuclear and epithelial cells obtained from children who had adenoid tissue resected as part of standard treatment [32].

Moller et al demonstrated extensive invasion of lung explants from patients with end-stage lung disease (including COPD and cystic fibrosis) with H. influenza; the organism was present in the epithelium, the submucosa of the bronchi, the bronchioles, the interstitium, and the alveolar epithelium in over half of the subjects [33]. Dromann et al have demonstrated the presence of H. influenzae in lung tissue in 40 % of subjects in all stages of COPD [34]. Another study found very high levels of intracellular NTHi present in subjects who had had exacerbations of COPD [35].

Haemophilus influenzae has a number of mechanisms that allow it to persist in the human host. The ability to be able to invade into lung tissue may be particularly important in the pathogenesis of this bacterium. Key mechanisms are summarized in Table 1.

Table 1 Table 1 Important pathogenic features of nontypeable Haemophilus influenzae Go to: Immune response NTHi is present in the nasopharynx of most healthy adults but only causes clinical disease in a minority of subjects it infects. Therefore the nature of the respiratory tract immune/inflammatory response may be very important in the pathogenesis of this bacterium. NTHi causes strong stimulation of both innate and adaptive immunity.

Innate immune responses

Innate immunity serves as the first-line of defense against infection and is comprised of both structural and cellular defenses.

Structural defenses have a number of components including cough, barrier function and the mucociliary apparatus. Impairment of mucocilary function as occurs in cystic fibrosis and immotile cilia syndrome, is associated with lung H. influenzae infection.

The cellular innate immune response is a rapidly evolving area. It is mediated by neutrophils and particularly macrophages, which recognize bacterial pathogens by toll-like receptors (TLRs). The outer membrane proteins of H. influenzae such as P2 and P6 strongly activate innate immunity. P6 activates macrophages to produce interleukin (IL) 8 and tumor necrosis factor alpha (TNF-α) [36,37]. P6 is also important for the migration of dendritic cells. NTHi lysate (i.e. killed/inactivated bacteria) upregulates the production of nuclear transcription factor-kappa β (NF-κβ), which is a key driver of inflammation [38]. Epidermal growth factor receptor (EGFR) has also recently been shown to be important in NF-κβ activation [39].

The NF-κβ pathway is principally activated through toll-like receptors (TLRs) which function as pattern receptors (e.g. for bacterial motifs). Activation of TLRs by bacterial motifs drives an inflammatory response, which is designed to clear infection. Both TLR-2 and TLR-4 have been shown to be important in immune responses to NTHi [40,41].

TLR-4 primarily recognizes lipopolysaccharide (LPS) a component of the gram-negative cell wall. NTHi has lipooligosaccharide (LOS), which is very similar to LPS but lacks O-antigen units. Activation of TLR4 results in the production of a variety of inflammatory mediators by the macrophage, which have a key role in initiating innate immunity.

TLR-2 is also expressed on the surface of innate and immune cells. Outer membrane P6 activates TLR-2 with up-regulation of NF-κβ and the production of inflammatory mediators.

Adaptive immune responses

The adaptive immune response develops after the innate response and is particularly important in chronic infectious disease. It is primarily mediated by B lymphocytes (humoral immunity) and T lymphocytes (cellular immunity).

Studies have demonstrated that the great majority of healthy subjects and those with chronic airways disease have strong antibody responses to NTHi. It has also been shown that complement is bactericidal for NTHi. Antibody causes activation of the terminal attack complex of complement and this is very effective in killing NTHi [42]. This mechanism may explain why NTHi is primarily a mucosal pathogen that rarely spreads beyond the respiratory tract in contrast to the typeable forms such as Hib which are protected by the tough polysaccharide capsule and frequently cause systemic disease. Hypogammaglobulinaemia has been shown to be an important risk factor for systemic infection with NTHi.

T cells have a key role in the protection against intracellular infection. Both T helper (Th) cells and cytotoxic T (CTL) cells secrete cytokines (which drive inflammation) and produce cytotoxic mediators. The ability of T cells to proliferate to NTHi stimulation influences clinical disease. Patients with obstructive airways disease have deficient T-cell responses to NTHi when compared to healthy controls specifically; 1), decreased Th1 cell function, particularly production of interferon gamma (IFN-γ) & CD40 ligand (CD40L) [43] and 2), decreased CTL cell function, particularly production of IFN-γ [44]. In addition macrophage killing of NTHi is enhanced by the addition of IFN-γ & CD40L [45]. A recent study has confirmed that Th1 cell responses to NTHi in COPD are deficient [46] and TLRs appear to be important in this mechanism.

There are two other important situations, which have potentially significant effects on the immune response to NTHi in the lung. They are cigarette smoke and viral infections. These will now be discussed in more detail.

Effect of cigarette smoking on the lung immune response

Cigarette smoking is the key risk factor for the development of COPD and has a significant effect on the lung immune response [47]. Smoking directly damages the respiratory epithelium and inhibits mucociliary function [48,49]. It has a wide variety of effects on alveolar macrophages, dendritic cells and lymphocytes which inhibit the ability of the lung to clear infection [47]. Smoking has been found to be associated with increased lung inflammation following challenge with NTHi [50].

=Interaction with viral infection

Interaction between viral infection and Haemophilus influenzae

In clinical practice there is a frequent association between viral infection and bacterial infection. In the context of H. influenzae infection the two best-described associations have been with 1), epidemic influenza and 2), rhinovirus.

Exacerbations of COPD are frequently associated with combined viral and bacterial infection and the most common co-infection is the combination of rhinovirus and H. influenzae, which is also associated with increased severity of exacerbations [51,52]. There is preliminary evidence from laboratory studies that rhinovirus may interact with NTHi. Rhinovirus suppresses host macrophage interleukin 1 responses to NTHi [53] and enhances migration of NTHi across epithelial cells [54].

A major cause of death in epidemic viral influenza is secondary bacterial lung infection. Outbreaks are associated with different bacterial infections. The “Spanish flu” outbreak following World War 1 was associated with a high incidence of H. influenzae infection [55,56]. Viral influenza causes significant damage to the respiratory epithelial barrier and this may be an important mechanism of secondary bacterial infection.

Clinical presentation

CLINICAL MANIFESTATIONS OF H. INFLUENZAE INFECTIONS Upper respiratory tract infections. (i) Otitis media in children. Otitis media is the most frequently diagnosed bacterial illness in young children requiring office or clinic visits in the United States. Acute otitis media is an inflammation of the middle ear (the cavity between the eardrum and the inner ear), and episodes are characterized by fever, ear pain, and, in severe cases, discharge from the ear. The gold standard for an etiologic diagnosis, culture of middle ear fluid, requires tympanocentesis, which is an invasive procedure and is thus not routinely performed. However, based on results of middle ear fluid cultures obtained by tympanocentesis as part of clinical trials, H. influenzae is one of the most common causes of otitis media, accounting for 25 to 35% of episodes of acute otitis media (32).

Children who have 4 or more episodes of acute otitis media in a year or who experience at least 8 months of middle ear effusion in a year are defined as otitis prone. Up to 10% of children in the United States are otitis prone. Nasopharyngeal colonization by H. influenzae early in life is associated with the otitis-prone condition. The most common cause of recurrent otitis media is nontypeable H. influenzae (25). In developing countries, complications, including chronic suppurative otitis media, are frequent. Globally, up to 330 million people suffer from recurrent and chronic and otitis media (http://www.who.int/pbd/deafness/activities/hearing_care/otitis_media.pdf).

Since 2000, most infants in the United States have received the 7-valent pneumococcal conjugate vaccine. Widespread administration of this vaccine has resulted in a dramatic reduction in nasopharyngeal colonization by the 7 pneumococcal serotypes included in the vaccine. This has been accompanied by several important changes in the distribution of bacterial pathogens that cause otitis media: (i) a reduction in pneumococcal vaccine serotypes, (ii) a relative increase in pneumococcal nonvaccine serotypes, and (iii) a relative increase in H. influenzae and Moraxella catarrhalis causing nasopharyngeal colonization and otitis media (4, 9, 36). Two new pneumococcal conjugate vaccines have recently been approved for use, and these include a 13-valent vaccine in the United States and a 10-valent vaccine in Europe in which pneumococcal polysaccharides are conjugated to protein D, a surface protein of H. influenzae (34). These vaccines will have a significant impact on pathogens of otitis media, patterns of nasopharyngeal colonization, and the populations of bacterial strains that circulate in communities. Careful surveillance will be critical to track inevitable but not entirely predictable changes.

(ii) Sinusitis. Sinusitis is a complication of viral upper respiratory tract infection, estimated to occur following ∼7% of episodes in children and less often in adults. Samples obtained by sinus puncture or endoscopy reveal that H. influenzae is an important pathogen in both acute and chronic sinusitis (6, 7, 11). Pneumococcal conjugate vaccines appear to have resulted in a relative increase in H. influenzae as a cause of sinusitis (7). Because treatment of most sinusitis is empirical, H. influenzae should be considered when making antibiotic choices in patients with the clinical picture of bacterial sinusitis.

(iii) Conjunctivitis. Nontypeable H. influenzae causes conjunctivitis in children, the elderly, health care workers, and immunocompromised hosts. Person-to-person transmission, particularly in day care centers and nursing homes, plays an important role in the spread of infection in these settings (14, 47).

Lower respiratory tract infections. (i) Exacerbations in adults with COPD. Chronic obstructive pulmonary disease (COPD) afflicts ∼24 million Americans and is the fourth most common cause of death in the United States and in the world. The course of the disease is characterized by intermittent worsening or exacerbations that are associated with enormous morbidity, including missed work time, office and clinic visits, emergency room visits, hospital admissions, and, in the most severe cases, respiratory failure necessitating mechanical ventilation. The role of bacterial infection in COPD has long been a source of confusion and controversy, but more recently, investigation is beginning to elucidate the important role of bacteria in the course and pathogenesis of COPD (39). The best estimates are that approximately 30% of exacerbations are caused by viral infection and ∼50% are caused by bacterial infection. The presence of a bacterial pathogen in the sputum of an adult with COPD does not establish the etiology of an exacerbation because the same pathogens colonize COPD patient airways during clinically stable periods. Several lines of evidence, as follows, support the conclusions that bacteria cause exacerbations of COPD and that nontypeable H. influenzae is the most common bacterial cause of exacerbations of COPD.

Nontypeable H. influenzae is isolated from the lower airways of adults with exacerbations of COPD when the distal airways are sampled using bronchoscopy and the protected specimen brush. Acquisition of a new strain of H. influenzae is highly associated with the onset of exacerbation of COPD (31, 38). Adults with COPD who experience exacerbations associated with new strain acquisitions of H. influenzae develop strain-specific serum antibody responses (40). Acquisition of H. influenzae causes increased airway inflammation, a hallmark of exacerbations of COPD. H. influenzae and other bacterial pathogens also play a more subtle role in the pathogenesis of COPD. In contrast to the sterile airways of a healthy lung, respiratory pathogens are present in the airways of 25 to 50% of clinically stable adults with COPD, with H. influenzae as the most common. These pathogens release highly active antigens that induce enhanced airway inflammation, which is a characteristic feature of COPD.

(ii) Pneumonia. Precise data on the role of H. influenzae as a cause of pneumonia in children and adults are lacking. In the pre-Hib conjugate vaccine era, Hib was thought to be a predominant cause in children. The role of nontypeable H. influenzae in childhood pneumonia remains unclear, but several indirect lines of evidence, including its high prevalence in nasopharyngeal colonization studies, its demonstrated pathogenic potential in otitis media and preliminary evidence of involvement in bronchitis, suggest that nontypeable strains play some role in lower respiratory tract infection in children (15).

Nontypeable H. influenzae causes community-acquired pneumonia in adults, particularly in the elderly and in patients with COPD, HIV, and cystic fibrosis. Hence, antibiotics with activity against H. influenzae are recommended by Infectious Diseases Society of America guidelines when considering empirical treatment of community-acquired pneumonia in adults (26).

(iii) Infections in cystic fibrosis. Colonization and infection by Pseudomonas aeruginosa in patients with cystic fibrosis are often preceded by infection with nontypeable H. influenzae or Staphylococcus aureus early in the course of the disease. The presence of H. influenzae biofilms in the airways of children with cystic fibrosis suggests that biofilms may be important in early lung injury, facilitating later infection by P. aeruginosa (42).

Invasive infections. H. influenzae occasionally causes invasive infections in the post-Hib vaccine era; most such infections are caused by nontypeable strains. The highest incidence is in children less than 1 year old and in the elderly. The most common presentations are bacteremia and meningitis. Many but not all have some associated predisposing conditions, including immunosuppression, HIV infection, or a respiratory disorder. Nontypeable strains that cause invasive infection are genetically diverse.

Infections by other encapsulated strains have disease manifestations similar to those of type b strains, causing predominantly meningitis and invasive disease.