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By Kevin C. Hazen, PhD, Professor of Pathology and Director of Clinical Microbiology, Duke University Health System

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Culture is microbial growth on or in a nutritional solid or liquid medium; increased numbers of organisms simplify identification. Culture also facilitates testing of antimicrobial susceptibility.

Communication with the laboratory is essential. Although most specimens are placed on general purpose media (eg, blood or chocolate agar), some pathogens require inclusion of specific nutrients and inhibitors or other special conditions (see Table: Selective Media for Isolation of Common Bacteria); if one of these pathogens is suspected or if the patient has been taking antimicrobials, the laboratory should be advised. The specimen’s source is reported so that the laboratory can differentiate pathogens from site-specific normal flora.

Selective Media for Isolation of Common Bacteria


Preferred Medium

Bacteroides sp

Kanamycin-vancomycin laked blood agar

Bacteroides fragilis

Bacteroides bile-esculin (with gentamicin and bile)

Bordetella pertussis

Bordet-Gengou agar plus methicillin or cephalexin

Regan-Lowe cephalexin agar

Horse blood–charcoal agar

Burkholderia cepacia

Pseudomonas cepacia agar

Campylobacter jejuni or C. coli

Campylobacter-selective agars (eg, cefoperazone-vancomycin agar)

Corynebacterium diphtheriae

Tinsdale agar

Cystine-tellurite blood agar

Löffler coagulated serum medium

Escherichia coli or enterohemorrhagic pathogens (Shiga toxin producers, including O157-H7)

MacConkey-sorbitol agar

Francisella tularensis

Blood- or chocolate-cystine agar

Legionella sp

Buffered charcoal yeast extract agar

Leptospira sp

Fletcher or Stuart medium with rabbit serum or Leptospira medium with bovine serum albumin-Tween 80

Neisseria gonorrhoeae or N. meningitidis

Modified Thayer-Martin agar

New York City agar

Salmonella and Shigella sp

May grow on standard MacConkey or eosin-methylene blue

Alternative: Hektoen or xylose-lysine-desoxycholate, Salmonella-Shigella agar, gram-negative or selenite enrichment broth

Vibrio sp

Thiosulfate-citrate-bile salts–sucrose agar

Yersinia sp

Cefsulodin-Irgasan-novobiocin agar

Specimen collection

Specimen collection is important. For diagnosis of infectious disease, the rule of thumb is sample where the infection is. For lesions, the leading edge, not the center, should be sampled.

Use of swabs is discouraged. However, if a swab is used, a flocked swab is preferred because it can recover more specimen. Swabs used for molecular assays (see Nucleic Acid–Based Identification Methods for Infectious Disease) must be compatible for the specific molecular assay for which they are intended. The wrong type of swab can produce false-negative results. Wooden-shafted swabs are toxic to some viruses. Cotton-tipped swabs are toxic for some bacteria, including chlamydiae.

Blood cultures require decontamination and disinfection of the skin (eg, povidone iodine swab, allowed to dry, removed with 70% alcohol). Multiple samples, each from a different site are generally used; they are taken nearly simultaneously with fever spikes if possible. Normal flora of skin and mucous membranes that grows in only a single blood sample is usually interpreted as contamination. If a blood specimen is obtained from a central line, a peripheral blood specimen should also be obtained to help differentiate systemic bacteremia from catheter infection. Cultures from infected catheters generally turn positive more quickly and contain more organisms than simultaneously drawn peripheral blood cultures. Some fungi, particularly molds (eg, Aspergillus spp), usually cannot be cultured from blood.

The specimen must be transported rapidly, in the correct medium, and in conditions that limit growth of any potentially contaminating normal flora. For accurate quantification of the pathogen, additional pathogen growth must be prevented; specimens should be transported to the laboratory immediately or, if transport is delayed, refrigerated (in most cases).

Special considerations for culture

Certain cultures have special considerations.

Anaerobic bacteria should not be cultured from sites where they are normal flora because differentiation of pathogens from normal flora may be impossible. Specimens must be shielded from air, which can be difficult. For swab specimens, anaerobic transport media are available. However, fluid specimens (eg, abscess contents) are superior to swab specimens for recovery of anaerobic bacteria. Fluid specimens should be collected with a syringe from which all air was expressed (to minimize contact of the specimen with oxygen) and sent to a laboratory in the syringe (capped without the needle) or transferred to an anaerobic transport vial.

Mycobacteria are difficult to culture. Specimens containing normal flora (eg, sputum) must first be decontaminated and concentrated. Mycobacterium tuberculosis and some other mycobacteria grow slowly. Growth of M. tuberculosis is typically faster in liquid than in solid media; routine use of automated systems with liquid media can result in growth within 2 wk vs 4 wk on solid media such as Lowenstein-Jensen agar. In addition, few organisms may be present in a specimen. Multiple specimens from the same site may help maximize yield. Specimens should be allowed to grow for 8 wk before being discarded. If an atypical mycobacterium is suspected, the laboratory should be notified.

Viruses are generally cultured from swabs and tissue specimens usually transported in media that contain antibacterial and antifungal agents. Specimens are inoculated onto tissue cultures that support the suspected virus and inhibit all other microbes. Viruses that are highly labile (eg, varicella zoster) should be inoculated onto tissue cultures within 1 h of collection. Standard tissue cultures are most sensitive. Rapid tissue cultures (shell vials) may provide more rapid results. Some common viruses cannot be detected using routine culture methods and require alternative methods for diagnosis (see Table: Diagnostic Tests for Common Viruses That Do Not Grow in Routine Viral Cultures), as for the following:

  • Enzyme immunoassay for Epstein-Barr virus, hepatitis B and E viruses, HIV, and human T-lymphotropic virus

  • Serologic tests for hepatitis A and D viruses

  • Nucleic acid–based methods for HIV

Diagnostic Tests for Common Viruses That Do Not Grow in Routine Viral Cultures

Common Conditions


Diagnostic Tests

Acute febrile illness, meningoencephalitis

Alphavirus, flaviviruses, bunyaviruses (eg, St. Louis encephalitis virus, La Crosse encephalitis virus)

EIA, nucleic acid–based methods


Rotaviruses, caliciviruses (noroviruses), astroviruses

EM or IEM,nucleic acid–based methods

Infectious mononucleosis

Epstein-Barr virus

EIA, nucleic acid–based methods

Hemorrhagic fevers, lymphocytic choriomeningitis

Filoviruses, arenaviruses (eg, Lassa fever, Ebola virus)

EM, nucleic acid–based methods


Hepatitis A, hepatitis D

Serologic testing, nucleic acid–based methods

Hepatitis B, hepatitis E

EIA, nucleic acid–based methods

Hepatitis C, hepatitis G

Nucleic acid–based methods, EIA

Roseola, Kaposi sarcoma, disseminated infections

Herpesviruses 6, 7, 8

Nucleic acid–based methods, EIA



Nucleic acid–based methods, EIA, Western blot

Condylomata acuminata, genital skin cancer

Human papillomaviruses

Nucleic-acid–based methods, EIA

Fifth disease

Human parvovirus B19

Nucleic acid–based methods, EIA

Adult T-cell leukemia

Human T-lymphotropic virus

EIA, nucleic acid–based methods

Progressive multifocal leukoencephalopathy, kidney infection

Polyoma viruses (JC and BK)

Nucleic acid–based methods

Smallpox, monkeypox, vaccinia, molluscum contagiosum


Nucleic acid–based methods, EM, culture depending on virus


Rabies virus

EM, IFA, nucleic acid–based methods


Rubella virus

EIA, IFA, nucleic acid–based methods

EIA = enzyme immunoassay; EM = electron microscopy; IEM = immunoelectron microscopy; IFA = immunofluorescence assay.

Fungi specimens obtained from nonsterile sites must be inoculated onto media containing antibacterial agents. Specimens should be allowed to grow for 3 to 4 wk before being discarded.

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