Infections of the Lower Respiratory System

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Infections of the Lower Respiratory System

Objectives

1. Define the trachea, bronchi, bronchioles, and alveoli, and explain the anatomic structure of the lower respiratory system.

2. List the most common etiologic agents responsible for lower respiratory disease and pneumonia in patients of various ages and categories: children <5 years of age, school-age children, young adults, older adults, and immunocompromised patients.

3. Describe the virulence factors found in bacteria and viruses associated with infection of the lower respiratory tract.

4. List the four possible routes of transmission or dissemination within the body that allow organisms to cause an infection in the lungs.

5. Name the most important decision for physicians regarding the treatment of pneumonia in older individuals, and list the three-step process used to guide them in this decision.

6. List the most prevalent cause of community-acquired pneumonia in adults.

7. Differentiate between community-acquired and hospital-acquired pneumonia.

8. State the factors anaerobic bacteria possess that enhance their ability to produce disease; explain how these anaerobes gain entrance to the lungs.

9. Define Lukens trap, and explain the type of patient or specimen associated with the method.

10. Describe the difference between early-onset or late-onset hospital- or ventilator-associated pneumonia.

11. List the etiologic agent of lung infections identified in cystic fibrosis patients.

12. Name the organisms most often associated with pneumo-opportunistic infection in HIV-positive individuals.

13. Explain the mechanisms that, because of the bacterial production of toxins, enable microorganisms to produce respiratory-associated disease.

14. Explain how the host immune system can contribute to microorganism growth in the respiratory disease process.

15. Explain why Mycobacterium tuberculosis is a classic representative of an intracellular pathogen.

16. Describe specimens collected for respiratory infections including determination of specimen quality and rejection criteria for the following: sputum, induced sputum, endotracheal suction, pleural fluid, bronchoalveolar lavage, bronchial washing, and bronchial brush sample.

17. Explain how the microbiologist would test for the less common causes of respiratory infection, including Pneumocystis jiroveci, Legionella spp., Chlamydophila pneumonia, Bordetella pertussis, Mycoplasma pneumonia, and Norcardia.

General Considerations

Anatomy

The respiratory tract can be divided into two major areas: the upper respiratory tract consists of all structures above the larynx, whereas the lower respiratory tract follows airflow below the larynx through the trachea to the bronchi and bronchioles and then into the alveolar spaces where gas exchange occurs (Figure 69-1). The respiratory and gastrointestinal tracts are the two major connections between the interior of the body and the outside environment. The respiratory tract is the pathway through which the body acquires fresh oxygen and removes unneeded carbon dioxide. It begins with the nasal and oral passages, which humidify inspired air, and extends past the nasopharynx and oropharynx to the trachea and then into the lungs. The trachea divides into bronchi, which subdivide into bronchioles, the smallest branches that terminate in the alveoli. Some 300 million alveoli are estimated to be present in the lungs; these are the primary microscopic gas exchange structures of the respiratory tract.

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Figure 69-1 Anatomy of the respiratory tract, including upper and lower respiratory tract regions.

Familiarization with the anatomic structure of the thoracic cavity ensures proper specimen collection from various sites in the lower respiratory tract for processing by the laboratory. The thoracic cavity, which contains the heart and lungs, has three partitions separated from one another by pleura (see Figure 69-1). The lungs occupy the right and left pleural cavities, whereas the mediastinum (space between the lungs) is occupied mainly by the esophagus, trachea, large blood vessels, and heart.

Pathogenesis of the Respiratory Tract: Basic Concepts

Microorganisms primarily cause disease by a limited number of pathogenic mechanisms (see Chapter 3). Because these mechanisms relate to respiratory tract infections, they are discussed briefly. Encounters between the human body and microorganisms occur many times each day. However, establishment of infection after such contact tends to be the exception rather than the rule. Whether an organism is successful in establishing an infection depends not only on the organism’s ability to cause disease (pathogenicity) but also on the human host’s ability to prevent the infection.

Host Factors

The human host has several mechanisms that nonspecifically protect the respiratory tract from infection: the nasal hairs, convoluted passages, and the mucous lining of the nasal turbinates; secretory IgA and nonspecific antibacterial substances (lysozyme) in respiratory secretions; the cilia and mucous lining of the trachea; and reflexes such as coughing, sneezing, and swallowing. These mechanisms prevent foreign objects or organisms from entering the bronchi and gaining access to the lungs, which remain sterile in the healthy host. Aspiration of minor amounts of oropharyngeal material, as occurs often during sleep, plays an important role in the pathogenesis of many types of pneumonia. Once particles escape the mucociliary sweeping activity and enter the alveoli, alveolar macrophages ingest them and carry them to the lymphatics.

In addition to these nonspecific host defenses, normal flora of the nasopharynx and oropharynx help prevent colonization by pathogenic organisms of the upper respiratory tract. Normal bacterial flora prevent the colonization by pathogens by competing for the same space and nutrients as well as production of bacteriocins and metabolic products that are toxic to invading organisms. Some of the bacteria that can be isolated as part of the indigenous flora of healthy hosts, as well as many species that may cause disease under certain circumstances and are often isolated from the respiratory tracts of healthy persons, are listed in Box 69-1. Under certain circumstances and for unknown reasons, these colonizing organisms can cause disease—perhaps because of previous damage by a viral infection, loss of some host immunity, or physical damage to the respiratory epithelium (e.g., from smoking). Differentiation of normal flora of the respiratory tract is important for determining the importance of an isolate in the clinical laboratory. Colonization does not always represent an infection. It is important to differentiate colonization from infection based on the specimen source, number of organisms present, and presence or quantity of white blood cells. (Organisms isolated from normally sterile sites in the respiratory tract by sterile methods that avoid contamination with normal flora should be definitively identified and reported to the clinician.)

Box 69-1   Organisms Present in the Nasopharynx and Oropharynx of Healthy Humans

Microorganism Factors

Organisms possess traits or produce products that promote colonization and subsequent infection in the host. The virulence, or disease-producing capability of an organism, depends on several factors including adherence, production of toxins, amount of growth or proliferation, tissue damage, avoiding the host immune response, and ability to disseminate.

Adherence.

For any organism to cause disease, it must first gain a foothold within the respiratory tract to grow to sufficient numbers to produce symptoms. Therefore, most etiologic agents of respiratory tract disease must first adhere to the mucosa of the respiratory tract. The presence of normal flora and the overall state of the host affect the ability of microorganisms to adhere. Surviving or growing on host tissue without causing overt harmful effects is termed colonization. Except for those microorganisms inhaled directly into the lungs, all etiologic agents of disease must first colonize the respiratory tract before they can cause harm.

Streptococcus pyogenes possess specific adherence factors such as fimbriae comprised of molecules such as lipoteichoic acids and M proteins. These molecules appear as a thin layer of fuzz surrounding the bacteria. Staphylococcus aureus and certain viridans streptococci are other bacteria that posses these lipoteichoic acid adherence complexes. Many gram-negative bacteria (which do not have lipoteichoic acids), including Enterobacteriaceae, Legionella spp., Pseudomonas spp., Bordetella pertussis, and Haemophilus spp., also adhere by means of proteinaceous finger-like surface fimbriae. Viruses possess either a hemagglutinin (influenza and parainfluenza viruses) or other proteins that mediate their epithelial attachment.

Toxins.

Certain microorganisms are almost always considered to be etiologic agents of disease if they are present in any numbers in the respiratory tract because they possess virulence factors that are expressed in every host. These organisms are listed in Box 69-2. The production of extracellular toxin was one of the first pathogenic mechanisms discovered among bacteria. Corynebacterium diphtheriae is a classic example of a bacterium that produces disease through the action of an extracellular toxin. Once the organism colonizes the upper respiratory epithelium, it produces a toxin that is disseminated systemically, adhering preferentially to central nervous system cells and muscle cells of the heart. Systemic disease is characterized by myocarditis, peripheral neuritis, and local disease that can lead to respiratory distress. Growth of C. diphtheriae causes necrosis and sloughing of the epithelial mucosa, producing a “diphtheritic (pseudo) membrane,” which may extend from the anterior nasal mucosa to the bronchi or may be limited to any area between—most often the tonsillar and peritonsillar areas. The membrane may cause sore throat and interfere with respiration and swallowing. Although nontoxic strains of C. diphtheriae can cause local disease, it is much milder than disease associated with toxigenic strains.

Box 69-2   Respiratory Tract Pathogens

Some strains of Pseudomonas aeruginosa produce a toxin similar to diphtheria toxin. Whether this toxin actually contributes to the pathogenesis of respiratory tract infection with P. aeruginosa has not been established. Bordetella pertussis, the agent of whooping cough, also produces toxins. The role of these toxins in production of disease is not clear. They may act to inhibit the activity of phagocytic cells or to damage cells of the respiratory tract. Staphylococcus aureus and beta-hemolytic streptococci produce extracellular enzymes capable of damaging host cells or tissues. Extracellular products of staphylococci aid in the production of tissue necrosis and the destruction of phagocytic cells and contribute to the abscess formation associated with infection caused by this organism. Although S. aureus can be recovered from throat specimens, it has not been proved to cause pharyngitis. Enzymes of streptococci, including hyaluronidase, allow rapid dissemination of the bacteria. Many other etiologic agents of respiratory tract infection also produce extracellular enzymes and toxins.

Microorganism Growth.

In addition to adherence and toxin production, pathogens cause disease by merely growing in host tissue, interfering with normal tissue function, and attracting host immune effectors, such as neutrophils and macrophages. Once these cells begin to attack the invading pathogens and repair the damaged host tissue, an expanding reaction ensues with more nonspecific and immunologic factors being attracted to the area, increasing the amount of host tissue damage. Respiratory viral infections usually progress in this manner, as do many types of pneumonias, such as those caused by Streptococcus pneumoniae, S. pyogenes, Staphylococcus aureus, Haemophilus influenzae, Neisseria meningitidis, Moraxella catarrhalis, Mycoplasma pneumoniae, Mycobacterium tuberculosis, and most gram-negative bacilli.

Avoiding the Host Response.

Another virulence mechanism present in various respiratory tract pathogens is the ability to evade host defense mechanisms. S. pneumoniae, N. meningitidis, H. influenzae, Klebsiella pneumoniae, mucoid P. aeruginosa, Cryptococcus neoformans, and others possess polysaccharide capsules that serve both to prevent engulfment by phagocytic host cells and to protect somatic antigens from being exposed to host immunoglobulins. The capsular material is produced in such abundance by certain bacteria, such as pneumococci, that soluble polysaccharide antigen particles can bind host antibodies, blocking them from serving as opsonins. Vaccine consisting of capsular antigens provides host protection to infection, indicating that the capsular polysaccharide is a major virulence mechanism of H. influenzae, S. pneumoniae, and N. meningitidis.

Some respiratory pathogens evade the host immune system by multiplying within host cells. Chlamydia trachomatis, Chlamydia psittaci, Chlamydia pneumoniae, and all viruses replicate within host cells. They have evolved methods for being taken in by the “nonprofessional” phagocytic cells of the host to where they thrive within the intracellular environment. Once within these cells, the organism is protected from host humoral immune factors and other phagocytic cells. This protection lasts until the host cell becomes sufficiently damaged that the organism is then recognized as foreign by the host and is attacked. A second group of organisms that cause respiratory tract disease comprises organisms capable of survival within phagocytic host cells (usually macrophages). Once inside the phagocytic cell, these respiratory tract pathogens are able to multiply. Legionella, Pneumocystis jiroveci (Pneumocystis carinii), and Histoplasma capsulatum are some of the more common intracellular pathogens.

Mycobacterium tuberculosis is the classic representative of an intracellular pathogen. In primary tuberculosis, the organism is carried to an alveolus in a droplet nucleus, a tiny aerosol particle containing tubercle bacilli. Once phagocytized by alveolar macrophages, organisms are carried to the nearest lymph node, usually in the hilar or other mediastinal chains. In the lymph node, the organisms slowly multiply within macrophages. Ultimately, M. tuberculosis destroys the macrophage and is subsequently taken up by other phagocytic cells. Tubercle bacilli multiply to a critical mass within the protected environment of the macrophages, which are prevented from accomplishing phagosome-lysosome fusion capable of destroying the bacteria. Having reached a critical mass, the organisms spill out of the destroyed macrophages, through the lymphatics, and into the bloodstream, producing mycobacteremia and carrying tubercle bacilli to many parts of the body. In most cases, the host immune system reacts sufficiently at this point to kill the bacilli; however, a small reservoir of live bacteria may be left in areas of normally high oxygen concentration, such as the apical (top) portion of the lung. These bacilli are walled off, and years later, an insult to the host, either immunologic or physical, may cause breakdown of the focus of latent tubercle bacilli, allowing active multiplication and disease (secondary tuberculosis). In certain patients with primary immune defects, the initial bacteremia seeds bacteria throughout a compromised host, leading to disseminated or miliary tuberculosis. Growth of the bacteria within host macrophages and histiocytes in the lung causes an influx of more effector cells, including lymphocytes, neutrophils, and histiocytes, eventually resulting in granuloma formation, then tissue destruction and cavity formation. The lesion consists of a semisolid, amorphous tissue mass resembling semisoft cheese, from which it received the name caseating necrosis (death of cells or tissues). The infection can extend into bronchioles and bronchi from which bacteria are disseminated via respiratory secretions and coughing. Aerosolized droplets are produced by coughing and contain organisms that are inhaled by the next susceptible host. Other portions of the patient’s lungs may become infected as well through aspiration (inhalation of a fluid or solid).

Diseases of the Lower Respiratory Tract

Bronchitis

Acute

Acute bronchitis is characterized by acute inflammation of the tracheobronchial tree. This condition may be part of, or preceded by, an upper respiratory tract infection such as influenza (the “flu”) or the common cold. Most infections occur during the winter when acute respiratory tract infections are common.

The pathogenesis of acute bronchitis has no specific documented etiology but appears to be a mixture of viral cytopathic events and a response by the host immune system. Regardless of the cause, the protective functions of the bronchial epithelium are disturbed and excessive fluid accumulates in the bronchi. Depending on the etiology, destruction of the bronchial epithelium may be either extensive (e.g., influenza virus) or minimal (e.g., rhinovirus colds).

Clinically, bronchitis is characterized by cough, variable fever, and sputum production. Sputum (pus from the lungs) is often clear at the onset but may become purulent as the illness persists. Bronchitis may manifest as croup (a clinical condition marked by a barking cough or hoarseness).

The value of microbiologic studies to determine the cause of acute bronchitis in otherwise healthy individuals has not been established. Acute bronchitis is caused by viral agents, such as influenza and respiratory syncytial virus (RSV). The bacterium Bordetella pertussis is often associated with bronchitis in infants and preschool children (Table 69-1). The best specimen for diagnosis of pertussis is a deep nasopharyngeal specimen collected with a calcium alginate swab (see Chapter 37).

TABLE 69-1

Major Causes of Acute Bronchitis

Bacteria Viruses
Bordetella pertussis,
B. parapertussis,
Mycoplasma pneumoniae,
Chlamydia pneumoniae		 Influenza virus, adenovirus, rhinovirus, coronavirus (other less common viruses: respiratory syncytial virus, human metapneumovirus, coxsackie A21 virus)

Chronic versus Acute

Chronic bronchitis is a common condition affecting about 10% to 25% of adults. This disease is defined by clinical symptoms in which excessive mucus production leads to coughing up sputum on most days during at least 3 consecutive months for more than 2 successive years. Cigarette smoking, infection, and inhalation of dust or fumes are important contributing factors. Acute bronchitis is not related to long-term etiologies causing damage to the lungs, but is typically a result of an infectious process.

Patients with chronic bronchitis can suffer from acute flare-ups of infection, but determination of the cause of the infection is difficult. Potentially pathogenic bacteria, such as nonencapsulated strains of Haemophilus influenzae, Streptococcus pneumoniae, and Moraxella catarrhalis, are frequently cultured from the bronchi of these patients. Because of chronic colonization, it is difficult to incriminate one of these organisms as the specific cause of an acute infection in patients with chronic bronchitis. Although the role of bacteria in acute infections in these patients is questionable, viruses are frequent causes.

Bronchiolitis

Bronchiolitis, the inflammation of the smaller diameter bronchiolar epithelial surfaces, is an acute viral lower respiratory tract infection that primarily occurs during the first 2 years of life. Characteristic clinical manifestations include an acute onset of wheezing and hyperinflation as well as cough, rhinorrhea (runny nose), tachypnea (rapid breathing), and respiratory distress. The disease is primarily caused by viruses including a recently discovered virus, human metapneumovirus. RSV accounts for 40% to 80% of cases of bronchiolitis and demonstrates a marked seasonality; the etiologic agents of bronchiolitis are listed in Box 69-3. Like other viral infections, bronchiolitis shows a marked seasonality in temperate climates with a yearly increase in cases during winter to early spring.

Box 69-3   Viral Agents That Cause Bronchiolitis

Initially, the virus replicates in the epithelium of the upper respiratory tract, but in the infant it rapidly spreads to the lower tract airways. Early inflammation of the bronchial epithelium progresses to necrosis. Symptoms such as wheezing may be related to the type of inflammatory response to the virus as well as other host factors. For the most part, patients are managed based on clinical parameters, with the laboratory having a role in cases that require hospitalization; a specific viral etiology can be identified in a large number of infants by viral isolation from respiratory secretions, preferably from a nasal wash (see Chapter 65).

Pneumonia

Pneumonia (inflammation of the lower respiratory tract involving the lung’s airways and supporting structures) is a major cause of illness and death. There are two major categories of pneumonias: those considered community-acquired pneumonia (patients are believed to have acquired their infection outside the hospital setting) and those including hospital- or ventilator-associated (patients are believed to have acquired their infection within the hospital setting, usually at least 2 days following admission) or health care–associated pneumonia (affects only patients hospitalized in an acute care hospital for 2 or more days within 90 days of infection from a long-term care facility, or patients who have received recent intravenous antibiotic therapy, chemotherapy, or wound care within 30 days of the current infection, or who have attended a hospital or hemolysis clinic). Nevertheless, once a microorganism has successfully invaded the lung, disease can follow affecting the alveolar spaces and their supporting structure, the interstitium, and the terminal bronchioles.

Pathogenesis

Organisms can cause infection of the lung by four possible routes: by upper airway colonization or infection that subsequently extends into the lung, by aspiration of organisms (thereby avoiding the upper airway defenses), by inhalation of airborne droplets containing the organism, or by seeding of the lung via the blood from a distant site of infection. Viruses cause primary infections of the respiratory tract, as well as inhibit host defenses that, in turn, can lead to a secondary bacterial infection. For example, viruses may destroy respiratory epithelium and disrupt normal ciliary activity. Presumably, the growth of viruses in host cells disrupts the function of the latter and encourages the influx of nonspecific immune effector cells exacerbating the damage. Damage to host epithelial tissue by virus infection is known to predispose patients to secondary bacterial infection.

Aspiration of oropharyngeal contents is important in the pathogenesis of many types of pneumonia. Aspiration may occur during a loss of consciousness such as during anesthesia or a seizure, or after alcohol or drug abuse, but other individuals, particularly geriatric patients, may also develop aspiration pneumonia. Neurologic disease or esophageal pathology and periodontal disease or gingivitis are other important risk factors. Aided by gravity and often by loss of some host nonspecific protective mechanisms, organisms reach lung tissue, where they multiply and attract host inflammatory cells. Other mechanisms include inhalation of aerosolized material and hematogenous seeding. The buildup of cell debris and fluid contributes to the loss of lung function and thus to the pathology.

Furthermore, regarding the pathogenesis of hospital-associated, health care–associated, and ventilator-associated pneumonias, health care devices, the environment, and the transfer between the patient and staff or other patients can serve as sources of pathogens causing pneumonia. The primary routes for bacterial entry into the lower respiratory tract are by aspiration of oropharyngeal organisms or leakage of secretions containing bacteria around an endotracheal tube. For these reasons, intubation and mechanical ventilation significantly increase the risk of pneumonia (6- to 21-fold). In addition, bacterial and viral biofilm in the endotracheal tube with subsequent spread to distal airways may be important in the pathogenesis of ventilator-associated pneumonia.

Clinical Manifestations

The symptoms suggestive of pneumonia include fever, chills, chest pain, and cough. In the past, pneumonias were classified into two major groups: (1) typical or acute pneumonias (e.g., Streptococcus pneumoniae) and (2) atypical pneumonias, based on whether the cough was productive or nonproductive of mucoid sputum. However, analysis of symptoms of pneumonia caused by the atypical pneumonia pathogens (Mycoplasma pneumoniae, Legionella pneumophila, and Chlamydophila pneumoniae) has revealed no significant differences from those symptoms of patients with typical bacterial pneumonias. Because of this overlap in symptoms, it is important to consider all possible etiologies associated with the patient’s clinical presentation.

Some patients with pneumonia exhibit no signs or symptoms related to their respiratory tract (i.e., some only have fever). Therefore, physical examination of the patient, chest radiograph findings, patient history, and clinical laboratory findings are important. In addition to respiratory symptoms, 10% to 30% of patients with pneumonia complain of headache, nausea, vomiting, abdominal pain, diarrhea, and myalgias.

Epidemiology/Etiologic Agents

As previously mentioned, there are two major categories of pneumonias: those considered community-acquired pneumonias and hospital-, ventilator-, or health care–associated pneumonias. Because the epidemiology and etiologies can differ, these two categories are discussed separately. Pneumonia in the immunocompromised patient is addressed separately in this chapter. Emerging viral infections associated with severe acute respiratory syndrome (SARS) and influenza outbreaks (H1N1) are typically associated with upper respiratory infections but may lead to serious lower respiratory infections in the young, elderly, or immunocompromised patient. See Chapter 66 for detailed information related to these emerging viral infectious diseases and diagnostic recommendations.

Community-Acquired Pneumonia.

In the United States, pneumonia is the sixth leading cause of death and the number one cause of death from infectious diseases. It is estimated that as many as 2 million to 3 million cases of community-acquired pneumonia occur annually, and roughly one fifth of these require hospitalization; 45,000 pneumonia-related deaths occur in the United States each year. The etiology of acute pneumonias is strongly dependent on age. More than 80% of pneumonias in infants and children are caused by viruses, compared to less than 10% to 20% of pneumonias in adults.

Children.

Community-acquired pneumonia in children is a common and potentially serious infection. The annual incidence of pneumonia in children younger than 5 years of age is 34 to 40 cases per 1000 in Europe and North America. Determining the cause of pneumonia is challenging because the lungs are rarely sampled directly and sputum is difficult to obtain from children. Among previously healthy patients 2 months to 5 years old, RSV, human metapneumovirus, parainfluenza, influenza, and adenoviruses are the most common etiologic agents of lower respiratory tract disease. Children suffer less commonly from bacterial pneumonia, usually caused by H. influenzae, S. pneumoniae, or S. aureus. Neonates may acquire lower respiratory tract infections with C. trachomatis or P. jiroveci (which likely indicates an immature immune system or an underlying immune defect).

M. pneumoniae and C. pneumoniae are the most common causes of bacterial pneumonia in school-age children (5-14 years of age). The four most common causes of community-acquired viral pneumonia in children include influenza, RSV, parainfluenza, and adenovirus. The agents associated with nosocomial outbreaks in children include the influenza virus, RSV, and adenovirus. Mixed viral and bacterial infections have been documented in 35% of patients, with the majority of these (81%) being mixed viral-bacterial infections. In addition, the time of onset of hospital- or ventilator-associated pneumonia is an important epidemiologic variable and risk factor: early-onset pneumonia (defined as occurring within the first 4 days of hospitalization), usually carries a better prognosis, being more likely to be caused by antibiotic-sensitive bacteria, whereas late-onset pneumonia (5 days or more) is more likely to be caused by multidrug-resistant organisms and is associated with increased patient morbidity and mortality.

Young Adults.

The most common etiologic agent of lower respiratory tract infection among adults younger than 30 years of age is Mycoplasma pneumoniae, which is transmitted via close contact. Contact with secretions seems to be more important than inhalation of aerosols for transmission and infection. After contact with respiratory mucosa, Mycoplasma are able to adhere to and colonize respiratory mucosal cells. Both a protein adherence factor and gliding motility determine virulence. Mycoplasma attach to the cilia of respiratory mucosal cells; once there, they multiply and destroy ciliary function. Attachment and cytotoxins produced by the organisms induce cell damage. Chlamydia pneumoniae is the third most common agent of lower respiratory tract infection in young adults, following mycoplasmas and influenza viruses; it also affects older individuals. Chlamydia spp., intracellular pathogens capable of disrupting cellular function and causing respiratory disease, are similar to viral pathogens.

The epidemiology and treatment of community-acquired and hospital-acquired pneumonia have changed dramatically as a result of improvements in diagnostics, antimicrobial therapy, and supportive care modalities. The changes in the organization of health care has made the distinction between community-acquired and hospital-acquired pneumonia less clear. However, pneumonia still remains an important cause of morbidity and mortality in elderly patients. The American Thoracic Society and the Infectious Disease Society of America guidelines have suggested that patients who have been hospitalized in the last 90 days, reside in a nursing home or long-term care facility, or have had a recent intravenous antibiotic therapy or hemodialysis, be classified as a patient with health care-associated pneumonia (HCAP). Patients with health care-associated pneumonia have a higher incidence of cardiopulmonary and neurodegenerative diseases, cancer, chronic kidney disease, chronic obstructive pulmonary disease, and immunosuppression than elderly patients with community-acquired pneumonia. Both populations become infested with various organisms. The organisms most frequently responsible for community-acquired pneumonia include S. pneumoniae, H. influenzae, M. pneumoniae, C. pneumoniae, M. catarrhalis, and Legionella spp. Factors that contribute to the onset include decreased mucociliary function, decreased cough reflex, decreased level of consciousness, periodontal disease, and decreased general mobility. Health care-associated patients have been found to be more frequently colonized with gram-negative bacilli and other multidrug resistant pathogens, perhaps because of poor oral hygiene, decreased saliva, or decreased epithelial cell turnover. The microorganisms associated with these infections, in addition to those previously mentioned, may include methicillin-resistant S. aureus (MRSA), Pseudomonas aeruginosa, a variety of Enterobacteriaceae, Acinetobacter spp., anaerobic bacteria, carbapenamase-resistant Klebsiella pneumonia, and extended spectrum beta-lactamase resistant Enterobacteriaceae (ESBLS). According to the Infectious Diseases Society of America (IDSA), the decision to hospitalize a patient or to treat him or her as an outpatient is possibly the single most important clinical decision made by physicians during the course of illness. This decision in turn impacts the subsequent site of treatment (home, hospital, or intensive care unit), intensity of laboratory evaluation, antibiotic therapy, and cost. Thus, the IDSA has developed management guidelines for community-acquired pneumonia in adults based on a three-step process: (1) assessment of preexisting conditions that might compromise safety of home care, (2) quantification of short-term mortality (referred to as the pneumonia port severity index [PSI] and based on a prediction rule derived from more than 14,000 patients) with subsequent assignment of patients to five risk classes (classes I through V), and (3) clinical judgment usually require hospitalization. The PSI, however, is not useful for patients in nursing homes or other health care facilities. It is therefore essential to properly assess the severity of the disease in both cases of community-acquired and health care associated pneumonia in elderly patients that clearly includes the three major management guidelines as outlined by the IDSA.

Pneumonia secondary to aspiration of gastric or oral sections is common and occurs in the community setting.

Pneumonia secondary to aspiration of gastric or oral secretions is common and occurs in the community setting. The most common agents include the oral anaerobes such as black-pigmented Prevotella and Porphyromonas spp., Prevotella oris, P. buccae, P. disiens, Bacteroides gracilis, fusobacteria, and anaerobic or microaerophilic streptococci. The anaerobic agents possess many factors, such as extracellular enzymes and capsules enhancing their ability to produce disease. It is their presence, however, in an abnormal site within the host producing lowered oxidation-reduction potential secondary to tissue damage that contributes to their pathogenicity. Staphylococcus aureus, various Enterobacteriaceae, and Pseudomonas may also be acquired by aspiration; Haemophilus influenzae, Legionella spp., Acinetobacter, Moraxella catarrhalis, Chlamydia pneumoniae, meningococci, and other agents may also be implicated. Pnuemonia is the leading cause of death among patients with nosocomial infections (hospital- ventilator- and health care-associated) (as high as 50%) mortality among patients in intensive care units. Some of these pneumonias are secondary to sepsis, and some are related to contaminated inhalation therapy equipment, particularly for intubated patients. Hospitalized patients or long-term care patients may experience asymptomatic colonization of the upper airway and result in aspiration of microorganisms into the lower respiratory tract. In addition to those organisms previously listed, these patients are more prone to infections with the multi-drug resistant strains of bacteria (ESBLS and MRSA) including Providencia stuartii, Morganella morganii, E. coli, Proteus mirabilis, K. pneumoniae, Enterobacter spp., and Staphylococcus aureus.

Adults (Viral pneumonia).

Adults may suffer from an estimated 100 million cases annually of community-acquired viral pneumonia cased by influenza, adenovirus, enteroviruses (coxsackieviruses and rhinoviruses), coronaviruses, human metapneumovirus, parainfluenza, varicella, rubeola or RSV, particularly during epidemics. Influenza associated viral pneumonia poses an increased risk for pregnant women of approximately 4-9 times greater than the general public, with the greatest risk associated with the third trimester. RSV is considered the third most common cause of community-acquired pneumoniae with 78% of the deaths occurring in patients over the age of 65. Similarly to RSV, human metapneumovirus has been associated with outbreaks in long-term care facilities. Following viral pneumonia, secondary bacterial disease caused by beta-hemolytic streptococci, S. aureus, M. catarrhalis, H. influenzae, and Chlamydia pneumoniae. Other agents may be considered depending on the geographic location and clinical presentation are viruses in the Hantavirus group, the most common of which is sin nombre virus as well as severe acute respiratory syndrome (SARS). (See Chapter 65.)

Of these agents, influenza virus, RSV and adenovirus have been implicated in nosocomial outbreaks. The time of onset of hospital- or ventilator-associated pneumonia is an important epidemiologic variable and risk factor; early onset pneumonia (defined as occurring within the first 4 days of hospitalization.

Adults (Fungal pneumonia).

Unusual causes of acute lower respiratory tract infection in adults include Actinomyces and Nocardia spp. Other agents may rarely be recovered from sputum and include the agents of plague, tularemia, melioidosis (Burkholderia pseudomallei), Brucella, Salmonella, Coxiella burnetii (Q fever), Bacillus anthracis, Pasteurella multocida, and certain parasitic agents such as Paragonimus westermani, Entamoeba histolytica, Ascaris lumbricoides, and Strongyloides spp. (the latter may cause fatal disease in immunosuppressed patients). A high index of suspicion by the clinician is usually a prerequisite to a diagnosis of parasitic pneumonia in the United States. Psittacosis should be ruled out as a cause of acute lower respiratory tract infection in patients who have had recent contact with birds. Among the fungal etiologies, Histoplasma capsulatum, Blastomyces dermatitidis, Paracoccidioides brasiliensis, Coccidioides immitis, Cryptococcus neoformans, and, occasionally, Aspergillus fumigatus may cause acute pneumonia. Therefore, occupational history and history of exposure to animals are important in suggesting specific potential infectious agents.

Chronic Lower Respiratory Tract Infections.

Mycobacterium tuberculosis is the most likely etiologic agent of chronic lower respiratory tract infection, but fungal infection and anaerobic pleuropulmonary infection may also run a subacute or chronic course. Mycobacteria other than M. tuberculosis may also cause such disease, particularly M. avium complex and M. kansasii. Although possible causes of acute, community-acquired lower respiratory tract infections, fungi and parasites are more commonly isolated from patients with chronic disease. Actinomyces and Nocardia may also be associated with gradual onset of symptoms. Actinomyces is usually associated with an infection of the pleura or chest wall, and Nocardia may be isolated along with an infection caused by M. tuberculosis. The pathogenesis of many of the infections caused by agents of chronic lower respiratory tract disease is characterized by the requirement for breakdown of cell-mediated immunity in the host or the ability of these agents to avoid being destroyed by host cell-mediated immune mechanisms. This may be caused by an effect on macrophages, the ability to mask foreign antigens, sheer size, or some other factor, allowing microbes to grow within host tissues without eliciting an overwhelming local immune reaction.

Cystic fibrosis (CF) is a genetic disorder that leads to persistent bacterial infection in the lung, causing airway wall damage and chronic obstructive lung disease. Eventually, a combination of airway secretions and damage leads to poor gas exchange in the lungs, cardiac malfunction, and subsequent death. Patients with CF may present as young adults with chronic respiratory tract disease or, more commonly, as children with gastrointestinal problems and stunted growth. Staphylococcus aureus is the most prevalent opportunistic bacterial pathogen infecting 55% of children 0–9 years of age with CF, with Pseudomonas aeruginosa the most prevalent (81%) in older children. A very mucoid Pseudomonas, characterized by production of copious amounts of extracellular capsular polysaccharide, can be isolated from the sputum of almost all patients with CF who are older than 18 years of age, becoming more prevalent with increasing age after 5 years. Even if CF has not been diagnosed, isolation of a mucoid Pseudomonas aeruginosa from sputum should alert the clinician to the possibility of underlying disease. Microbiologists should always report this unusual morphologic feature. In addition to mucoid Pseudomonas and Staphylococcus aureus, important pathogens in patients with CF are likely to harbor Haemophilus influenzae, Streptococcus pneumoniae, Stenotrophomonas maltophilia, Achromobacter xylosoxidans, Ralsotnia spp. Cupriavidus spp., Pandoraea spp., Escherichia coli, strains of Burkholderia cepacia complex, fast growing mycobacteria, RSV, influenza and fungi including Aspergillu, Scedosporium spp., and Exophiala dermatidis. In addition, due to the viscous mucous plugs associated with CF, several anaerobic organisms have been detected in the lungs of CF patients including Prevotella, Bifidobacterium, Veillonella, Peptostreptococcus and Fusobacterium. Using advanced diagnostic molecular methods, additional organisms have also been identified in chronic polymicrobial CF infections including viridans streptococci, Streptococcus constellatus, Streptococcus intermedius and Streptococcus anginosus.

Lung abscess is usually a complication of acute or chronic pneumonia. In these circumstances, organisms infecting the lung cause localized destruction of the lung parenchyma (functional elements of the lung). Symptoms associated with lung abscess are similar to those of acute and chronic pneumonia, except symptoms fail to resolve with treatment.

Immunocompromised Patients.

Patients with Neoplasms.

Patients with cancer are at high risk to become infected because of either granulocytopenia or other defects in phagocytic defenses, cellular or humoral immune dysfunction, damage to mucosal surfaces and the skin, and various medical procedures such as blood product transfusion. In these patients, the nature of the malignancy often determines the etiology (Table 69-2) and pneumonia is a frequent clinical manifestation.

TABLE 69-4

Examples of Infectious Agents Frequently Associated with Certain Malignancies

Malignancy (site and type of infections) Pathogens
Acute nonlymphocytic leukemia (pneumonia, oral lesions, cutaneous lesions, urinary tract infections, hepatitis, most often sepsis without obvious focus)

Acute lymphocytic leukemia (pneumonia, cutaneous lesions, pharyngitis, disseminated disease)

Lymphoma (disseminated disease, pneumonia, urinary tract infections, sepsis, cutaneous lesions)

Multiple myeloma (pneumonia, cutaneous lesions, sepsis)

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Transplant Recipients.

For successful organ transplantation, the recipient’s immune system must be suppressed. As a result, these patients are predisposed to infection. Regardless of the type of organ transplant (heart, renal, bone marrow, heart/lung, liver, pancreas), most infections occur within 4 months following transplantation. Major infections can occur within the first month but are usually associated with infections carried over from the pretransplant period. Pulmonary infections are of great importance in this patient population. Some of the most common causes of pneumonia include S. aureus Streptococcus pneumoniae, Haemophilus influenzae, Pneumocystis jiroveci, and cytomegalovirus. In addition, other organisms such as Cryptococcus neoformans, Aspergillus spp., Candida spp., Nocardia sp. and over, can cause life-threatening pulmonary infection.

HIV-Infected Patients.

Patients who are infected with human immunodeficiency virus (HIV) are at high risk for developing pneumonia. As discussed in the previous chapter, opportunistic infections as a result of severe immunodeficiency are a major cause of illness and death among these patients. In the United States, the most common opportunistic infection among patients with acquired immunodeficiency syndrome is Pneumocystis jiroveci pneumonia. Although P. jiroveci remains a major pulmonary pathogen, other organisms must be considered in this patient population, including Mycobacterium tuberculosis and Mycobacterium avium complex, as well as common bacterial pathogens such as Streptococcus pneumoniae and Haemophilus influenzae. In addition to these common pathogens, many other organisms can cause lower respiratory tract infections, including Nocardia spp., Rhodococcus equi (a gram-positive, aerobic, pleomorphic organism), and Legionella spp.

Laboratory Diagnosis of Lower Respiratory Tract Infections

Specimen Collection and Transport

Although rapid determination of the etiologic agent is of paramount importance in managing pneumonia, the responsible pathogen is not identified in as many as 50% of patients, despite extensive diagnostic testing. Unfortunately, no single test is capable of identifying all potential lower respiratory tract pathogens. Refer to Table 5-1 for an overview of the method used to collect, transport, and process specimens from the lower respiratory tract.

Sputum

Expectorated.

The examination of expectorated sputum has been the primary means of determining the causes of bacterial pneumonia. However, lower respiratory tract secretions will be contaminated with upper respiratory tract secretions, especially saliva, unless they are collected using an invasive technique. For this reason, sputum is among the least clinically relevant specimens received for culture in microbiology laboratories, even though it is one of the most numerous and time-consuming specimens.

Good sputum samples depend on thorough health care worker education and patient understanding throughout all phases of the collection process. Food should not have been ingested for 1 to 2 hours before expectoration and the mouth should be rinsed with saline or water just before expectoration. Patients should be instructed to provide a deep-coughed specimen. The material should be expelled into a sterile container, with an attempt to minimize contamination by saliva. Specimens should be transported to the laboratory immediately. Even a moderate amount of time at room temperature can result in the loss of viable infectious agents and the recovery of pathogens.

Induced.

Patients unable to produce sputum may be assisted by respiratory therapists, who use postural drainage and thoracic percussion to stimulate production of acceptable sputum. Before specimen collection, patients should brush the buccal mucosa, tongue, and gums with a wet toothbrush. As an alternative, an aerosol-induced specimen may be collected for the isolation of mycobacterial or fungal agents. Induced sputum is also recognized for its high diagnostic yield in cases of Pneumocystis jiroveci pneumonia. Aerosol-induced specimens are collected by allowing the patient to breathe aerosolized droplets, using an ultrasonic nebulizer containing 10% 0.85% NaCl or until a strong cough reflex is initiated. Lower respiratory secretions obtained in this way appear watery, resembling saliva, although they often contain material directly from alveolar spaces. These specimens are usually adequate for culture and should be accepted in the laboratory without prescreening. Obtaining such a specimen may obviate the need for a more invasive procedure, such as bronchoscopy or needle aspiration.

The gastric aspirate is used exclusively for isolation of acid-fast bacilli and may be collected from patients who are unable to produce sputum, particularly young children. Before the patient wakes up in the morning, a nasogastric tube is inserted into the stomach and contents are withdrawn (on the assumption that acid-fast bacilli from the respiratory tract were swallowed during the night and will be present in the stomach). The relative resistance of mycobacteria to acidity allows them to remain viable for a short period. Gastric aspirate specimens must be delivered to the laboratory immediately so that the acidity can be neutralized. Specimens can be neutralized and then transported if immediate delivery is not possible.

Endotracheal or Tracheostomy Suction Specimens

Patients with tracheostomies are unable to produce sputum in the normal fashion, but lower respiratory tract secretions can easily be collected in a Lukens trap (Figure 69-2). Tracheostomy aspirates or tracheostomy suction specimens should be treated as sputum by the laboratory. Patients with tracheostomies rapidly become colonized with gram-negative bacilli and other nosocomial pathogens. Such colonization per se is not clinically relevant, but these organisms may be aspirated into the lungs and cause pneumonia. Culture results should be correlated with clinical signs and symptoms.

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Figure 69-2 Tracheal secretions received in the laboratory in a Lukens trap.

Bronchoscopy.

Bronchoscopy specimens include bronchoalveolar lavage (BAL), bronchial washing, bronchial brushing, and transbronchial biopsies. The diagnosis of pneumonia, particularly in HIV-infected and other immunocompromised patients, often necessitates the use of more invasive procedures. Fiberoptic bronchoscopy has dramatically affected the evaluation and management of these infections. With this method, the bronchial mucosa can be directly visualized and collected for biopsy, and the lung tissue can be sent for transbronchial biopsy for the evaluation of lung cancer and other lung diseases. Although transbronchial biopsy is important, the procedure is often associated with significant complications such as bleeding. The sample should be transported in sterile 0.85% saline.

During bronchoscopy, physicians obtain bronchial washings or aspirates, bronchoalveolar lavage (BAL) samples, protected bronchial brush samples, or specimens for transbronchial biopsy. Bronchial washings or aspirates are collected using a small amount of sterile physiologic saline inserted into the bronchial tree and withdrawing the fluid. These specimens will be contaminated with upper respiratory tract flora such as viridans streptococci and Neisseria spp. Recovery of potentially pathogenic organisms from bronchial washings should be attempted.

A deep sampling of desquamated host cells and secretions can be collected through bronchoscopy and BAL. Lavages are especially suitable for detecting Pneumocystis cysts and fungal elements. During this procedure, a high volume of saline (100 to 300 mL) is infused into a lung segment through the bronchoscope to obtain cells and protein of the pulmonary interstitium and alveolar spaces. It is estimated that more than 1 million alveoli are sampled during this process. The value of this technique in conjunction with quantitative culture for the diagnosis of most major respiratory tract pathogens, including bacterial pneumonia, has been documented. Scientists have found significant correlation between acute bacterial pneumonia and greater than 103 to 104 bacterial colonies per milliliter of BAL fluid. BAL has been shown to be a safe and practical method for diagnosing opportunistic pulmonary infections in immunosuppressed patients. At bedside, nonbronchoscopic “mini BAL” using a Metras catheter has been introduced; typically 20 mL or less of saline is instilled.

Another type of respiratory specimen is obtained via a protected catheter bronchial brush as part of a bronchoscopy examination. Specimens obtained by this moderately invasive collection procedure are suited for microbiologic studies, particularly in aspiration pneumonia. Protected specimen brush bristles collect from 0.001 to 0.01 mL of material. An overview of the collection process is shown in Figure 69-3. Upon receipt, contents of the bronchial brush may be suspended in 1 mL of broth solution with vigorous vortexing and inoculated onto culture media using a 0.01-mL calibrated inoculating loop. Some researchers have indicated that specimens obtained via double-lumen–protected catheters are suitable for both anaerobic and aerobic cultures. Colony counts of greater than or equal to 1000 organisms per milliliter in the broth diluent (or 106/mL in the original specimen) have been considered to correlate with infection. All facets of the bronchoscopic procedure—such as order of sampling, use of anesthetic, and rapidity of plating—should be rigorously standardized.

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Figure 69-3 Overview for obtaining a protected catheter bronchial brush during a bronchoscopy examination. A, The bronchoscope is introduced into the nose and advanced through the nasopharyngeal passage into the trachea. The bronchoscope is then inserted into the lung area of interest. B, A small brush that holds 0.001 to 0.01 mL of secretions is placed within a double cannula. The end of the outermost tube or cannula is closed with a displaceable plug made of absorbable gel. The cannula is inserted to the proper area. C, Once in the correct area, the inner cannula is pushed out, dislodging the protective plug as it is extruded. D, The brush is then extended beyond the inner cannula, and the specimen is collected by “brushing” the involved area. The brush is withdrawn into the inner cannula, which is withdrawn into the outer cannula to prevent contamination by upper airway organisms as it is removed.

Transtracheal Aspirates.

Percutaneous transtracheal aspirates (TTAs) are obtained by inserting a small plastic catheter into the trachea via a needle previously inserted through the skin and cricothyroid membrane. This invasive procedure, although somewhat uncomfortable for the patient and not suitable for all patients (it cannot be used in uncooperative patients, in patients with bleeding tendency, or in patients with poor oxygenation), reduces the likelihood that a specimen will be contaminated by upper respiratory tract flora and diluted by added fluids, provided care is taken to keep the catheter from being coughed back up into the pharynx. Although this technique is rarely used, anaerobes, such as Actinomyces and those associated with aspiration pneumonia, can be isolated from TTA specimens.

Other Invasive Procedures.

When pleural empyema is present, thoracentesis may be used to obtain infected fluid for direct examination and culture. This constitutes an excellent specimen that accurately reflects the bacteriology of an associated pneumonia. Laboratory examination of such material is discussed in Chapter 77. Blood cultures, of course, should always be obtained from patients with pneumonia.

For patients with pneumonia, a thin needle aspiration of material from the involved area of the lung may be performed percutaneously. If no material is withdrawn into the syringe after the first try, approximately 3 mL of sterile saline can be injected and then withdrawn into the syringe. Patients with emphysema, uremia, thrombocytopenia, or pulmonary hypertension may be at increased risk of complication (primarily pneumothorax [air in the pleural space] or bleeding) from this procedure. The specimens obtained are very small in volume, and protection from aeration is usually impossible. This technique is more frequently used in children than in adults.

The most invasive procedure for obtaining respiratory tract specimens is the open lung biopsy. Performed by surgeons, this method is used to procure a wedge of lung tissue. Biopsy specimens are extremely helpful for diagnosing severe viral infections, such as herpes simplex pneumonia, for rapid diagnosis of Pneumocystis pneumonia, and for other hard-to-diagnose or life-threatening pneumonias. Ramifications of this and all other specimen collection techniques are discussed in Cumitech 7B, “Laboratory Diagnosis of Lower Respiratory Tract Infections.”

Specimen Processing

Direct Visual Examination

Lower respiratory tract specimens can be examined by direct wet preparation for parasites and special procedures for Pneumocystis. Fungal elements can be visualized under phase microscopy with 10% potassium hydroxide, under ultraviolet light with calcofluor white, or using periodic acid-Schiff–stained smears.

For most other evaluations, the specimen must be fixed and stained. Bacteria and yeasts can be recognized on Gram stain. One of the most important uses of the Gram stain, however, is to evaluate the quality of expectorated sputum received for routine bacteriologic culture. A portion of the specimen consisting of purulent material is chosen for the stain. The smear can be evaluated adequately even before it is stained, thus negating the need for Gram stain of specimens later judged unacceptable. An acceptable specimen yields fewer than 10 squamous epithelial cells per low-power field (100×). The number of white blood cells may not be relevant, because many patients are severely neutropenic and specimens from these patients will not show white blood cells on Gram stain examination. On the other hand, the presence of 25 or more polymorphonuclear leukocytes per 100× field, together with few squamous epithelial cells, implies an excellent specimen (Figure 69-4). Samples that contain predominantly upper respiratory tract material should be rejected. Previously, only expectorated sputa were suitable for rejection based on microscopic screening. However, endotracheal aspirates (ETAs) from mechanically ventilated adult patients can be screened by Gram stain. Criteria used to reject ETAs from adult patients include greater than 10 squamous epithelial cells per low-power field or no organisms seen under oil immersion (1000×). In Legionella pneumonia, sputum may be scant and watery, with few or no host cells. Such specimens may be positive by direct fluorescent antibody stain and culture, and they should not be subjected to screening procedures. Conversely, sputum from patients with CF should be screened. A throat swab is an acceptable specimen from patients with CF in selected clinical settings and should be processed in a similar manner as CF sputum. Staining of respiratory samples is useful and should be compared to culture results to reveal errors in procedures, specimen collection, and transport or specimen identification.

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Figure 69-4 Gram stain of sputum specimens. A, This specimen contains numerous polymorphonuclear leukocytes and no visible squamous epithelial cells, indicating that the specimen is acceptable for routine bacteriologic culture. B, This specimen contains numerous squamous epithelial cells and rare polymorphonuclear leukocytes, indicating an inadequate specimen for routine sputum culture.

Respiratory secretions may need to be concentrated before staining. The cytocentrifuge instrument has been used successfully for this purpose, concentrating the cellular material in an easily examined monolayer on a glass slide. As an alternative, specimens are centrifuged, and the sediment is used for visual examinations and cultures. For screening purposes, the presence of ciliated columnar bronchial epithelial cells, goblet cells, or pulmonary macrophages in specimens obtained by bronchoscopy or BAL indicates a specimen from the lower respiratory tract.

In addition to the Gram stain, respiratory specimens may be stained for acid-fast bacilli with either the classic Ziehl-Neelsen or the Kinyoun carbolfuchsin stain. Auramine or auramine-rhodamine is also used to detect acid-fast organisms. Because they are fluorescent, these stains fluorescent superior already here comment only are more sensitive than the carbolfuchsin formulas and are preferable for rapid screening. Slides may be restained with the classic stains directly over the fluorochrome stains as long as all of the immersion oil has been removed carefully with xylene. All of the acid-fast stains will reveal Cryptosporidium spp. if they are present in the respiratory tract, as may occur in immunosuppressed patients. These patients are often at risk of infection with P. jiroveci. Although the modified Gomori methenamine silver stain has been used traditionally to recognize Nocardia, Actinomyces, fungi, and parasites, it takes approximately 1 hour of the technologist’s time to perform, is technically demanding, and is not suitable as an emergency procedure. A fairly rapid stain, toluidine blue O, has been used in many laboratories with some success. Toluidine blue O stains Pneumocystis, Nocardia asteroides, and some fungi. A monoclonal antibody stain is the optimum stain for Pneumocystis (see Chapter 59) for less invasive specimens such as BAL and induced sputa.

Direct fluorescent antibody (DFA) staining has been used to detect Legionella spp. in lower respiratory tract specimens. Sputum, pleural fluid, aspirated material, and tissues are all suitable specimens. Because there are so many different serotypes of legionellae, polyclonal antibody reagents and a monoclonal antibody directed against all serotypes of Legionella pneumophila are used. Because of low sensitivity (50% to 75%), DFA results should not be relied on in lieu of culture. Rather, Legionella culture, DFA or urinary antigen, and serology should be performed for optimum sensitivity. See Chapter 35 for details regarding detection of Legionella spp.

Commercially available DFA reagents are also used to detect antigens of numerous viruses, including herpes simplex, cytomegalovirus, adenovirus, influenza viruses, and RSV (see Chapter 65). Commercial suppliers of reagents provide procedure information for each of these tests. Monoclonal and polyclonal fluorescent stains for Chlamydia trachomatis are available and may be useful for staining respiratory secretions of infants with pneumonia. A number of molecular amplification techniques (see Chapter 8) for the direct detection of respiratory pathogens have been described; however, the sensitivity and specificity of these assays vary greatly from one study to another. Amplification assays are also available for the direct detection of Mycobacterium tuberculosis on smear-positive specimens (see Chapter 43).

Rapid direct detection from respiratory samples is now available using nucleic acid-based methods. The xTAG Respiratory Viral Panel (RVP) (Luminex Corporation, Austin, TX), can be used for the simultaneous detection of influenza (four types), RSV, human metapneumovirus, and adenovirus from nasopharyngeal swabs. In addition, the FilmArray Respiratory Panel (BIOFIRE Diagnostics, Salt Lake City, UT ) is capable of detecting upper respiratory tract infections associated with coronavirus (four types), adenovirus, influenza (five types), rhinovirus, parainfluenza virus (four types), enterovirus, human metapneumovirus, RSV, Bordetella pertussis, Mycoplasma pneumoniae, and Chlamydophila pneumoniae in approximately 1 hour directly from patient samples. Smaller molecular panels are also available such as the real-time multiplex amplification kit for influenza A, B, and RSV (Hologic-Gen-Probe, San Diego, CA). All of the previously mentioned methods are FDA-approved. In addition to these, there are a variety of research-use-only and other molecular respiratory panels in clinical validation studies. It is important when considering the use of a molecular assay that the laboratory consider their patient population including severity of illness, immune status, and transplant histories.

Routine Culture

Most of the commonly sought etiologic agents of lower respiratory tract infection are isolated on routine media: 5% sheep blood agar, MacConkey agar for the isolation and differentiation of gram-negative bacilli, and chocolate agar for Haemophilus and Neisseria spp. Because of contaminating oral flora, sputum specimens, specimens obtained by bronchial washing and lavage, tracheal aspirates, and tracheostomy or endotracheal tube aspirates are not inoculated to enrichment broth or incubated anaerobically. Only specimens obtained by percutaneous aspiration (including transtracheal aspiration) and protected bronchial brush are suitable for anaerobic culture; the latter must be done quantitatively for proper interpretation (refer to prior discussion). Transtracheal and percutaneous lung aspiration material may be inoculated to enriched thioglycollate as well as to solid media. For suspected cases of Legionnaires’ disease, buffered charcoal-yeast extract (BCYE) agar and selective BCYE should be inoculated. Plates should be streaked in four quadrants to provide a basis for objective semiquantitation to define the amount of growth. After 24 to 48 hours of incubation, the numbers and types of colonies are recorded. For Legionella cultures, colonies form on the selective agar after 3 to 5 days at 35° C.

Sputum specimens from patients known to have CF should be inoculated to selective agar, such as specific chromagenic agar, for recovery of S. aureus and selective horse blood–bacitracin, incubated anaerobically and aerobically, for recovery of H. influenzae that may be obscured by the mucoid Pseudomonas on routine media. The use of a selective medium for B. cepacia, such as PC or OFPBL agars, is also necessary.

For interpretation of culture results on those specimens contaminated by normal oropharyngeal flora (e.g., expectorated and induced sputum, bronchial washings), growth of the predominant aerobic and facultative anaerobic bacteria is reported. To ensure optimum culture reporting, conditions must be well defined in terms of an objective grading system for streaked plates. Finally, the clinical significance of culture findings depends not only on standardized and appropriate laboratory methods but also on how specimens are collected and transported, other laboratory data, and the patient’s clinical presentation.

Numerous bacterial agents that cause lower respiratory tract infections are not detected by routine bacteriologic culture. Mycobacteria, Chlamydia, Nocardia, Bordetella pertussis, Legionella, and Mycoplasma pneumoniae require special procedures for detection; this also applies to viruses and fungi. Optimal recovery for Mycobacterium tuberculosis requires multiple specimens for acid-fast staining culture, and at least one sample for molecular testing as recommended by the Centers for Disease Control. Refer to the appropriate chapter section for more information regarding these organisms. Finally, one must keep in mind those potential agents for bioterrorist attack, such as Bacillus anthracis, Francisella tularensis, and Yersinia pestis, that might be recovered from respiratory specimens (see Chapter 80).

Chapter Review

1. Which of the following lung conditions are a major cause of illness and death?

2. What is the most frequent cause of community-acquired pneumonia in adults?

3. Psittacosis is a lower respiratory infection in humans caused by contact with what animal?

4. What organism may be recovered as a subacute illness in respiratory disease caused by M. tuberculosis?

5. All of the acid-fast stains will reveal which of the following organisms from respiratory specimens?

6. Which of the following stains is used to detect Legionella in lower respiratory specimens?

7. Which of the following media is used in addition to normal respiratory culture media in suspected cases of Legionnaires’ disease?

8. Which of the following organisms could potentially be isolated from a respiratory specimen and is not also considered a potential agent for bioterrorism?

9. Which of the following procedures has dramatically affected the evaluation and management of pneumonia, particularly in HIV-infected and immunocompromised patients?

10. True or False

_____ An acceptable sputum specimen when visualized by Gram stain must show significant white blood cells and fewer than 10 squamous epithelial cells per low-power field.

_____ Respiratory specimens first stained for acid-fast bacilli, with auramine or auramine-rhodamine, can be restained with the classic stains directly over the fluorochrome stain as long as all the immersion oil has been removed with xylene.

_____ A single culture is able to identify all potential pathogens that cause lower respiratory tract infections.

_____ Sputum is the most clinically relevant specimen received for culture in the microbiology laboratory for the diagnosis of respiratory infection.

_____ A moderate amount of time at room temperature will not affect the quality of a sputum sample.

_____ Patients with tracheostomies are unable to produce sputum in the normal fashion.

_____ The modified Gomori methenamine silver stain has been used traditionally to recognize Norcardia, Actinomyces, fungi, and parasites but is technically and time demanding.

_____ Pneumonia patients that are assessed to be risk classes II and III generally require hospitalization.

_____ The production of extracellular toxin was one of the first pathogenic mechanisms discovered in bacteria.

_____ The areas of the lungs most affected by pneumonia are the alveolar spaces and the interstitium, the supporting structure of the alveoli, and the terminal bronchioles.

11. Matching:

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12. Matching: Key Terms

Match the term with the appropriate definition.

13. Short Answer

1. Explain the benefits of using the Binax NOW S. pneumoniae urinary antigen test for presumptive diagnosis of pneumococcal pneumonia.

2. Why are enrichment broths or anaerobe plates not used in the culture of routine sputum specimens, specimens obtained by bronchial washing, and lavage tracheal aspirates?

3. What specimens are suited for anaerobic culture with regard to respiratory culture?

4. What characteristics do the following bacteria share that enable the organisms to resist engulfment by phagocytic host cells: S. pneumoniae, N. meningitides, H. influenzae, K. pneumoniae, P. aeruginosa, and Cryptococcus neoformans.

5. Define the pathogenesis of acute bronchitis.

6. When is thoracentesis used to obtain a sample for direct examination and culture?

7. What is an important epidemiologic variable and risk factor in hospital- or ventilator-associated pneumonia?

8. What virus infection effect may predispose patients to a secondary bacterial infection?

9. What hinders diagnosing the causative agent in pediatric pneumonia in children younger than 5 years of age?

Case Study 69-1

A 16-month-old boy was admitted with fever, lethargy, and trouble breathing. A diagnosis of pneumonia was made by physical examination. The child had recently been to Panama and was treated with ceftriaxone for cough and fever. His fever continued, despite treatment. On admission, he was given erythromycin therapy. Tracheal aspirate and blood cultures were obtained, but the respiratory specimen contained numerous epithelial cells and yielded normal respiratory flora on culture. A pleural aspirate and blood cultures were positive for Streptococcus pneumoniae, which was resistant to erythromycin and penicillin and intermediate in susceptibility to ceftriaxone. The patient was given high doses of ceftriaxone and vancomycin and responded to this therapy.

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