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Sputum cultures for the evaluation of bacterial pneumonia

Sputum cultures for the evaluation of bacterial pneumonia
Literature review current through: Jan 2024.
This topic last updated: Aug 15, 2023.

INTRODUCTION — Lower respiratory tract infections are common in the general population, occurring with increased frequency in older individuals and those with chronic diseases or compromised immune function. An etiologic diagnosis can be made by culture of respiratory tract secretions, by isolation of a compatible organism from blood or pleural fluid cultures, or by molecular methods.

While a positive blood or pleural fluid culture definitively identifies the pathogen, an organism growing from a respiratory specimen or detected by a molecular amplification method is not definitive proof that it is the etiologic agent. Many bacterial species are normal flora or colonizers of the respiratory tract and, although present in respiratory secretions, they may not be responsible for the clinical illness in an individual patient with pneumonia due to another cause. As a result, there is controversy about the diagnostic significance of many cultures or molecular results obtained from respiratory specimens. (See "Clinical evaluation and diagnostic testing for community-acquired pneumonia in adults" and "Epidemiology, pathogenesis, microbiology, and diagnosis of hospital-acquired and ventilator-associated pneumonia in adults" and "Clinical presentation and diagnostic evaluation of ventilator-associated pneumonia".)

A rational approach to specimen collection and the interpretation of results is required if clinically useful information is to be obtained [1]. The value and limitations of sputum cultures in patients with suspected bacterial pneumonia as well as the clinical indications for obtaining sputum cultures in such patients will be reviewed here.

Issues related to specimen transport to the laboratory, general approach to Gram stain and culture, and examples of clinical microbiology findings in patients with respiratory tract infections are discussed separately. (See "Microbiology specimen collection and transport" and "Approach to Gram stain and culture results in the microbiology laboratory".)

The approach to the evaluation of patients with possible tuberculosis, Pneumocystis jirovecii (formerly Pneumocystis carinii) pneumonia, invasive aspergillosis, influenza, and other respiratory viral infections is also discussed separately.

(See "Epidemiology, clinical presentation, and diagnosis of Pneumocystis pulmonary infection in patients with HIV".)

(See "Epidemiology, clinical manifestations, and diagnosis of Pneumocystis pneumonia in patients without HIV".)

(See "Epidemiology and clinical manifestations of invasive aspergillosis".)

(See "Seasonal influenza in adults: Clinical manifestations and diagnosis".)

(See "Parainfluenza viruses in adults".)

(See "Respiratory syncytial virus infection: Clinical features and diagnosis in infants and children".)

(See "Diagnosis, treatment, and prevention of adenovirus infection".)

(See "Diagnosis of pulmonary tuberculosis in adults".)

EXPECTORATED SPUTUM — Expectorated sputum is the most common lower respiratory tract specimen received by the microbiology laboratory. Expectorated sputum is often difficult to assess because many specimens are in fact mostly upper airway specimens and not sputum. The first and most important step in evaluation is the Gram stain. Gram stains are useful to:

Assess the suitability of the sputum specimen for further processing and interpretation

Predict a likely etiologic agent by identification of a predominant bacterial morphology in an adequate (purulent) specimen.

Several guidelines have been proposed to evaluate the quality of sputum samples. These guidelines have proposed different combinations and cutoffs of the minimum number of squamous epithelial cells (SECs) and/or polymorphonuclear leukocytes (PMNs) per low-power field (LPF; 10x objective) [2], but none of these parameters can be considered to be clearly superior. Our laboratory rejects all specimens with more than 10 SECs/LPF (picture 1) without considering the number of PMNs [3]. Rejection criteria have not been evaluated for Legionella, mycobacteria, or dimorphic fungi, and our laboratory accepts sputum specimens for diagnosis of these pathogens without screening for cellular content.

If a submitted specimen is rejected as unsuitable for culture, the clinician should be notified and a repeat specimen should be requested. If Gram stains are used for screening, the suitability of each sputum sample should be assessed as soon as possible after the sample arrives in the laboratory in order to minimize delays in acquiring another sample.

The diagnostic yield of the sputum Gram stain correlates directly with its quality. In one systematic review evaluating over 4500 patients with community-acquired pneumonia (CAP), bacterial pathogens were identified in 73 percent (95% credible interval [CrI] 26-96 percent) of good-quality specimens, but only 36 percent (95% CrI 22-53 percent) of all specimens regardless of quality. Sensitivity and specificity were highest for certain bacterial pathogens. For Streptococcus pneumoniae, the sensitivity was 0.69 (95% CI 0.56-0.99) and the specificity was 0.91 (95% CI 0.83-0.96); for Haemophilus influenzae, the sensitivity of 0.76 (95% CI 0.60-0.87) and a specificity of 0.97 (95% CI 0.91-0.99) [4].

While some purulent sputum specimens demonstrate multiple organisms rather than a single pathogen, they still can be useful (picture 2). For example, some fastidious organisms clearly seen on Gram stain may be overgrown by indigenous respiratory flora on solid media (eg, S. pneumoniae).

The sputum Gram stain has less utility in the diagnosis of atypical pneumonia. In general, patients with atypical pneumonia have mucoid sputum with abundant PMNs and few or no organisms on Gram stain. (See "Clinical manifestations and diagnosis of Legionella infection" and "Mycoplasma pneumoniae infection in adults".)

Collection of expectorated sputum for optimal yield — Up to one-third of patients with bacterial pneumonia may be unable to produce a sputum specimen, even under optimal conditions [5,6]. Approaches used to improve the quality of the specimen obtained include:

Obtaining the specimen prior to antibiotic treatment

Rinsing the mouth prior to expectoration

No food for one to two hours prior to expectoration

Inoculation of the culture media immediately after the specimen is obtained or immediately after prompt transport to the microbiology lab

When are sputum Gram stains and cultures indicated? — There is controversy about the utility of sputum specimens in CAP. The value of sputum examination and culture in patients with nosocomial pneumonia, especially ventilator-associated pneumonia, is more universally acknowledged.

Special stains of respiratory secretions may be performed for certain organisms when clinically indicated (eg, acid-fast stains for mycobacteria, direct fluorescent antibody staining for P. jirovecii). These are discussed in greater detail separately. (See "Clinical evaluation and diagnostic testing for community-acquired pneumonia in adults" and "Epidemiology, clinical manifestations, and diagnosis of Pneumocystis pneumonia in patients without HIV", section on 'Microscopy with staining'.)

Bronchitis — Sputum Gram stain and culture have no role in the evaluation of acute bronchitis in otherwise healthy individuals. Similarly, they are not indicated in the initial evaluation of patients with acute exacerbations of chronic obstructive pulmonary disease. (See "COPD exacerbations: Management" and "Acute bronchitis in adults", section on 'Microbiologic testing'.)

Community-acquired pneumonia — Generally, we obtain a sputum Gram stain and culture for most hospitalized patients with CAP, provided appropriate measures for collection, transport, and processing are in place so that a good quality sample can be obtained. Although the diagnostic yield is variable, identifying a pathogen allows for directed therapy and enhances knowledge of local epidemiology.

However, the approach to testing varies among experts and medical societies. As an example, the Infectious Diseases Society of America (IDSA) and the American Thoracic Society (ATS) guidelines recommend obtaining sputum Gram stain and culture in patients who have any of the following [7]:

Severe CAP, defined by ATS/IDSA criteria (see "Community-acquired pneumonia in adults: Assessing severity and determining the appropriate site of care")

History of colonization or past infection with methicillin-resistant Staphylococcus aureus (MRSA) or Pseudomonas aeruginosa

Current empiric treatment for MRSA, P. aeruginosa, or other multidrug-resistant gram-negative bacteria

Hospitalization with receipt of intravenous antibiotics in the past 90 days or other strong suspicion for MRSA or P. aeruginosa infection

Ideally, sputum studies should be obtained prior to starting treatment. An endotracheal sputum sample should be obtained from intubated patients.

Sputum tests are generally not recommended for outpatients, as empiric antibiotic therapy is generally successful and knowledge of the infecting pathogen does not usually improve outcomes. (See "Clinical evaluation and diagnostic testing for community-acquired pneumonia in adults", section on 'Sputum Gram stain and culture'.)

Hospital-acquired (nosocomial) pneumonia — Sputum Gram stain and culture are indicated for all patients with hospital-acquired pneumonia. (See "Clinical presentation and diagnostic evaluation of ventilator-associated pneumonia".)

Such specimens may consist of either expectorated sputum or suctioned secretions, depending upon whether the patient is intubated. One study correlated Gram stain and culture results for 504 consecutive sputum samples obtained by endotracheal suctioning [8]. A mean of 4.05 organisms were recovered from the 15 percent of specimens with >10 squamous epithelial cells per low-power field (SEC/LPF), compared with 2.35 organisms for specimens with fewer epithelial cells. The yield from culture was low (88 percent either no growth or normal oropharyngeal flora) when no organisms were visible by Gram stain. The authors of this study concluded that a laboratory that rejected endotracheal suctioned specimens with >10 SEC/LPF or no organisms under high power could generate considerable savings for the institution.

Sputum culture results cannot be interpreted in the absence of clinical data in patients suspected of having nosocomial pneumonia, especially those who are intubated. In patients suspected of having nosocomial pneumonia, a positive sputum culture in association with the typical signs and symptoms (fever, peripheral leukocytosis, deteriorating oxygenation, change in quantity or character of sputum, or a new infiltrate on chest radiograph) is suggestive of pneumonia. (See "Clinical presentation and diagnostic evaluation of ventilator-associated pneumonia".)

Purulent sputum and a culture that grows a predominant organism may be indicative of tracheobronchitis, rather than parenchymal infection, in a patient without the clinical findings described above. Furthermore, in such a patient, a nonpurulent specimen growing a predominant organism is not indicative of respiratory tract infection and should not be used as an indication to start antimicrobial therapy. Hospitalized patients become colonized rapidly with enteric gram-negative bacilli with a frequency that is dictated by the severity of their associated conditions and prior antibiotic exposure. It is often difficult or impossible to determine the clinical significance of these organisms.

SPUTUM CULTURES — Routine media used for the isolation and identification of respiratory pathogens include blood agar, chocolate agar, and MacConkey agar:

Blood agar supports the growth of gram-positive cocci and most gram-negative rods and is especially useful for evaluation of the colony morphology and hemolysis of streptococci.

Chocolate agar permits recovery of H. influenzae and other fastidious organisms that may grow less well on blood agar.

MacConkey agar is selective for gram-negative bacteria and allows further classification into lactose-positive or -negative organisms, based upon their ability to ferment lactose.

Specialized media is required for culture of atypical pathogens such as Legionella pneumophila. (See "Clinical manifestations and diagnosis of Legionella infection".)

The laboratory should be notified when respiratory specimens are sent from patients with cystic fibrosis so that special media and processing can be used to detect pathogens of particular interest in this patient population, such as Burkholderia cepacia.

Interpretation — Culture results are reported in a semiquantitative manner (1+ to 4+ in some laboratories; rare, few, moderate, or abundant in others). Most true pathogens are present in at least 3+ (moderate) amounts. When the same organism is detected from respiratory and blood cultures concurrently, it is strongly suggestive that the respiratory tract is the source of the bacteremia. However, the diagnosis of pneumonia must be based on clinical, rather than laboratory, criteria.

As stated above, sputum cultures may not yield a specific pathogen. Since the oropharynx and upper airway are virtually always colonized with indigenous flora, cultures of expectorated sputum will often grow mixed flora, even in the absence of bacterial lower respiratory tract infection (picture 3). For some organisms, the concentration in culture is important, and clinical correlation is critical.

The yield of sputum cultures is further diminished if the patient has received antibiotics prior to producing the specimen, which occurs frequently [5,6]. The greatest problem in this regard is with fastidious pathogens, such as S. pneumoniae and H. influenzae. The experienced technologist can select colonies with morphologies consistent with known respiratory pathogens, but their presence, especially in a mixed specimen, does not equate with causality.

Organisms can also colonize the respiratory tract, especially in patients with chronic illnesses or recent hospitalization. Thus, any positive sputum culture result must be interpreted in the context of the clinical setting. Some organisms are virtually never pulmonary pathogens, such as Candida spp, coagulase-negative staphylococci, and enterococci.

Specimens obtained via bronchoscopy — Bronchoscopy can be used to acquire samples of lower respiratory tract secretions in patients with suspected pneumonia in whom respiratory samples cannot be obtained by expectoration or in whom such samples have been nondiagnostic. Lower respiratory specimens can be obtained by bronchoalveolar lavage, routine brushing, washing, or protected specimen brushing (using a double-sheathed catheter that minimizes bacterial contamination). A variety of quantitative culture methods have been developed to differentiate infection from colonization and to identify pathogenic organisms in the fluid obtained, but none of these methods is completely reliable. The utility of quantitative cultures of specimens obtained by bronchoscopy has been highly variable in individual patients and in various studies. The role of bronchoscopic sampling and quantitative cultures, which have been best studied in patients with ventilator-associated pneumonia, are discussed separately. (See "Clinical presentation and diagnostic evaluation of ventilator-associated pneumonia", section on 'Noninvasive respiratory sampling'.)

When bronchoscopy is used, it is imperative that specimen handling be optimized, since fastidious organisms may not survive prolonged transport times. In addition, use of a diluent such as bacteriostatic saline in bronchoalveolar lavage or brush specimens can inhibit the growth of microorganisms, possibly leading to false-negative results [9].

Bronchoscopy for specimen collection is most useful for the diagnosis of infection due to Mycobacterium tuberculosis in patients with negative sputum studies, for P. jirovecii (formerly P. carinii) or other fungal or viral pathogens, or for establishing a noninfectious etiology such as malignancy [1]. (See "Flexible bronchoscopy in adults: Indications and contraindications" and "Clinical evaluation and diagnostic testing for community-acquired pneumonia in adults" and "Epidemiology, pathogenesis, microbiology, and diagnosis of hospital-acquired and ventilator-associated pneumonia in adults" and "Clinical presentation and diagnostic evaluation of ventilator-associated pneumonia" and "Epidemiology, clinical presentation, and diagnosis of Pneumocystis pulmonary infection in patients with HIV" and "Epidemiology, clinical manifestations, and diagnosis of Pneumocystis pneumonia in patients without HIV" and "Epidemiology and clinical manifestations of invasive aspergillosis" and "Diagnosis of pulmonary tuberculosis in adults" and "Seasonal influenza in adults: Clinical manifestations and diagnosis".)

SUMMARY

Lower respiratory tract infections are common in the general population, occurring with increased frequency in older individuals and those with chronic diseases or compromised immune function. A diagnosis is made by culture of respiratory tract secretions, by isolation of a compatible organism from blood or pleural fluid cultures, or by molecular methods. (See 'Introduction' above.)

While a positive blood or pleural fluid culture may definitively identify the pathogen, an organism growing from a respiratory specimen or detected by a molecular amplification assay is not definitive proof that it is the etiologic agent. Many bacterial species are normal flora or colonizers of the respiratory tract and, although present in respiratory secretions, they may not be responsible for the clinical illness in an individual patient with pneumonia due to another cause. (See 'Introduction' above.)

Expectorated sputum is the most common lower respiratory tract specimen received by the microbiology laboratory. Expectorated sputum is often difficult to assess because many specimens consist mostly of upper airway specimens and not sputum. The first and most important step in evaluation is the Gram stain. Gram stains are useful to (see 'Expectorated sputum' above):

Assess the suitability of the sputum specimen for further processing and interpretation

Predict a likely etiologic agent by identification of a predominant bacterial morphology in an adequate (purulent) specimen

Several guidelines have been proposed to evaluate the quality of sputum samples. These guidelines have proposed different combinations and cutoffs of the minimum number of squamous epithelial cells (SECs) and/or polymorphonuclear leukocytes (PMNs) per low-power field (LPF; 10x objective), but none of these parameters can be considered to be clearly superior. Our laboratory rejects all specimens with more than 10 SECs/LPF (picture 1) without considering the number of PMNs. (See 'Expectorated sputum' above.)

Sputum Gram stain and culture are indicated for all patients with hospital-acquired pneumonia and for certain patients with community-acquired pneumonia. Sputum Gram stain and culture have no role in the evaluation of acute bronchitis in otherwise healthy individuals. Similarly, they are not indicated in the initial evaluation of patients with acute exacerbations of chronic obstructive pulmonary disease. (See 'When are sputum Gram stains and cultures indicated?' above.)

Culture results are reported in a semiquantitative manner (1+ to 4+ in some laboratories, rare-few-moderate-abundant in others). Most true pathogens are present in at least 3+ (moderate) amounts. (See 'Interpretation' above.)

Any positive sputum culture result must be interpreted in the context of the clinical setting, since organisms can colonize the upper and/or lower respiratory tract. Some organisms are virtually never pulmonary pathogens, such as Candida spp, coagulase-negative staphylococci, and enterococci. (See 'Interpretation' above.)

Bronchoscopy can be used to acquire samples of lower respiratory tract secretions in patients with suspected pneumonia in whom respiratory samples cannot be obtained by expectoration or in whom such samples have been nondiagnostic. Bronchoscopy for specimen collection is most useful for the diagnosis of infection due to Mycobacterium tuberculosis in patients with negative sputum studies, for Pneumocystis jirovecii (formerly Pneumocystis carinii) or other fungal or viral pathogens, or for establishing a noninfectious etiology such as malignancy. (See 'Specimens obtained via bronchoscopy' above.)

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