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.

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 drain age 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 con tents 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 (Figure1). 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.

Fig1. 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 bronchos copy 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 2. Upon receipt, con tents 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.

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.

Fig2. 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 with drawn into the outer cannula to prevent contamination by upper airway organisms as it is removed. 

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.”

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مواضيع ذات صلة

tuberculosis testing (TB testing, Interferon gamma release assay [IGRA, T-Spot.TB], QuantiFERON-TB Gold [QFT, QFT-G, TB Gold test, TB blood test], Nucleic acid amplification for TB [NAAT], TB antibody)

thyrotropin-releasing hormone stimulation test (TRH stimulation test, Thyrotropin-releasing factor stimulation test [TRF stimulation test])

Laboratory Diagnosis of Lower Respiratory Tract Infections: Specimen Processing

thyroid scanning (Thyroid scintiscan)

thyroid fine needle aspiration biopsy (FNAB, Skinny needle thyroid biopsy, Fine-needle aspiration [FNA])

thoracentesis and pleural fluid analysis (Pleural tap)

The Role of Laboratory in Urgency: Altered State of Consciousness

Preoperative Laboratory Test

sputum culture and sensitivity (C&S, Culture and Gram stain)

sodium (Na), blood

small intestinal bacterial overgrowth test (SIBO test, Lactulose Breath Test)

Clinical Biochemistry of Exercise: The Benefits of Movement