Sputum Culture

Updated: May 26, 2022
Author: Majd Ibrahim, MD; Chief Editor: Daniela Hermelin, MD 

Reference Range

The most common pathogens detected with a sputum culture are bacteria such as Streptococcus pneumoniae, Haemophilus influenzae, Staphylococcus aureus, and Klebsiella species. Fungi are slow-growing eukaryotic organisms that can grow on living or nonliving organisms and are subdivided into molds and yeasts. Only a few of them grow in humans, and when they infect the respiratory system, they can cause serious infections. Overall, sputum specimens are observed for mucopurulent strands, leukocytes, and blood and culture results.[1, 2, 3, 4, 5]

The presence of normal upper respiratory tract flora should be expected in sputum culture. Normal respiratory flora include Neisseria catarrhalis, Candida albicans, diphtheroids, alpha-hemolytic streptococci, and some staphylococci.



A normal Gram stain of sputum contains polymorphonuclear leukocytes, alveolar macrophages, and a few squamous epithelial cells. The presence of normal flora does not rule out infection. Examination of a Gram-stained smear of the specimen frequently reveals whether the specimen is satisfactory or not. The quality of sputum samples is determined by the minimum number of squamous epithelial cells and polymorphonuclear leukocytes per low power field. An acceptable specimen has more than 25 leukocytes and fewer than 10 epithelial cells per low power field. An unacceptable sample can be misleading and should be rejected by the laboratory.

Culture of the sputum on blood agar frequently reveals characteristic colonies, and identification is made by various serologic or biochemical tests. Cultures of Mycoplasma are infrequently done; diagnosis is usually confirmed by a rise in antibody titer. If Legionella pneumonia is suspected, the organism can be cultured on charcoal-yeast agar, which contains the high concentrations of iron and sulfur required for growth.

If tuberculosis is suspected, an acid-fast stain should be performed immediately, and the sputum cultured on special media, which are incubated for at least 6 weeks.

In diagnosing aspiration pneumonia and lung abscesses, anaerobic cultures are important.


Collection and Panels

Equipment: Sterile, leak-proof container.

  • Collecting the first sample before any antibiotic or antimicrobial therapy is initiated is necessary.

  • Obtaining an early-morning expectorated specimen is most desirable. The first morning specimen is most concentrated and is less likely to be contaminated with saliva and nasopharyngeal secretions.

  • Before beginning collection, ask the client to rinse the mouth with plain water. This removes secretions and oral plaque, which may contaminate the sample.

  • Instruct the client to breathe deeply to stimulate coughing and expectoration. This loosens the secretions enough to expectorate.

  • Collect the expectorated sputum in a leak-proof sterile container. Refrigerate the container until processing takes place. Sterility is important for culture results. Refrigeration slows other bacterial growth.

  • Do not pool multiple samples in a 24-hr period. The client should be instructed to avoid adding saliva or nasopharyngeal secretions to the sputum sample. Avoids contamination of the sample.

  • Ask respiratory therapy personnel to assist the patient in obtaining an “aerosol-induced” specimen if the cough is not productive. Patients breathe aerosolized droplets of a sodium chloride-glycerin solution until a strong cough reflex is initiated. The specimen often appears watery but is in fact material directly from alveolar spaces. It should be noted on the requisition as being aerosol induced.

  • Cultures should be performed rapidly after collection, ideally within 2 hours; otherwise, the sample should be saved at 4ºC.




Sputum culture is used to diagnose pneumonia, bronchiectasis, bronchitis, or pulmonary abscess. It assists in the diagnosis of respiratory infections, as indicated by the presence or absence of organisms in culture.

The 2007 Infectious Diseases Society of America and American Thoracic Society (IDSA/ATS) consensus guidelines on community-acquired pneumonia (CAP) in adults recommend expectorated sputum specimens for hospitalized patients with signs and symptoms of pneumonia and any of the following conditions:[6]

  • Intensive care unit admission

  • Failure of outpatient antibiotic therapy

  • Cavitary lesions

  • Active alcohol abuse

  • Severe obstructive or structural lung disease

  • Positive urine antigen test for pneumococcus

  • Positive urine antigen test for Legionella (special culture media for Legionella needed)

  • Pleural effusion

Updated IDSA/ATS guidelines from 2019 state that, with regard to CAP, sputum culture is recommended not only for adult patients with severe disease but also for all adult inpatients who have received empirical treatment for methicillin-resistant S aureus and Pseudomonas aeruginosa.[7]

Sputum Gram stain and culture are indicated for all patients with hospital-acquired pneumonia.


See the list below:

  • Contamination with oral flora may invalidate results.

  • Specimen collection after antibiotic therapy has been initiated may result in inhibited or no growth of organisms.

  • Note any current antibiotic therapy on the laboratory slip.

A study by Huang et al indicated that with regard to predicting sputum culture results in pediatric patients with pneumonia, sputum Gram stains have high specificity and can therefore be used to guide antibiotic treatment choices. In terms of predicting gram-negative bacilli, gram-negative cocci, and gram-positive cocci, sputum Gram stains had specificities of 0.87, 0.98, and 0.87, respectively. Sensitivities were lower, at 0.45, 0.67, and 0.61, respectively.[8]