Blood Culture: Indications, Timing, and Volume

Blood culture is one of the most simple and commonly used investigations to establish the etiology of bloodstream infections (i.e. detection of bacteremia, sepsis, and fungemia). Early diagnosis and accurate identification of the bacteria or fungi causing bloodstream infections provides vital information to clinicians to start appropriate antimicrobial therapy. Providing timely and adequate antimicrobial therapy not only decreases infection-related mortality and cost but also reduces the risk of emergence/spread of drug resistance.

What is a blood culture?

A blood culture is a laboratory test in which blood, taken from the patient, is inoculated into bottles containing appropriate culture media to determine whether infection-causing microorganisms (bacteria or fungi) are present in the patient’s bloodstream.

Aims of blood culture:

  • To confirm presence of microorganisms in the bloodstream
  • To identify the etiology of the bloodstream infection
  • To guide appropriate antimicrobial therapy by providing susceptibility information of isolates

When should a blood culture be performed?

Blood cultures should always be requested when a bloodstream infection or sepsis is suspected. Some of the most common clinical symptoms in a patient which may lead to a suspicion of a bloodstream infection are:

  1. Undetermined fever ( ≥  38°C) or hypothermia ( ≤  36°C).
  2. Shock, chills, rigors
  3. Severe local infections (meningitis, endocarditis, pneumonia, pyelonephritis, intra-abdominal suppuration etc)

Blood cultures should be collected:

  • as soon as possible after the onset of clinical symptoms;
  • ideally, prior to the administration of antimicrobial therapy.

If the patient is already on antimicrobial therapy, recovery of microorganisms may be increased by collecting blood sample immediately before administering the next dose and by inoculating the blood into bottles containing specialized antimicrobial neutralization media.

What volume of blood should be collected?

Sufficient blood volume is critical for the successful recovery of organisms causing bloodstream infection. It has been found that most bacteremias in adults have a low number of colony-forming units (CFU) per milliliter (mL) of blood.

“More the blood volume cultured greater the chance of isolating pathogen causing bloodstream infection”.

Cumulative sensitivity of blood culture sets

Recommended blood-to-broth-ratio is 1:5 to 1:10, commercial blood culture systems may use a smaller blood-to-broth ratio (<1:5) due to the addition of sodium polyanetholesulfonate (SPS).


For an adult, the recommended volume of blood to be obtained per culture is 20-30 ml. Since each culture set includes an aerobic and an anaerobic bottle, each bottle should be inoculated with approximately 10 ml of blood (If anaerobic culture is not available in your settings, replacing it with an additional aerobic bottle ensures a culture of sufficient blood volume and increases the chance of recovery of pathogens).


For infants and children, the recommended blood volume should be based on the weight of the patient, and an aerobic bottle should be used, unless an anaerobic infection is suspected.

It is not safe to take large samples of blood from children, particularly infants. It is safe to obtain as much as 4-4.5% of known total blood volume for culture from infants and children. 

Specially formulated culture bottles are commercially available for use in children <2 years of age. They are specifically designed to maintain the usual blood-to-broth ratio (1:5 to 1:10) with smaller blood volumes, and have shown to improve microbial recovery.
For more information refer to table below:

Blood volumes suggested for cultures from infants and children
Blood volumes suggested for cultures from infants and children

How many blood culture sets should be collected?

Standards for the timing and number of blood cultures depends on the periodicity of microorganisms in the blood stream (random, intermittent, or continuous.

It is recommended to collect two, or preferably three blood culture sets for each septic episode. Since the pathogens may not be constantly present in the bloodstream (intermittent bacteremia), the sensitivity of a single blood culture set is always limited. Using single blood culture/bottle or set will result in an inadequate volume of blood cultured and a substantial number of bacteremias may be missed. It’s also difficult to confirm organism grown in a single blood culture bottle/set is contaminant or a true pathogen.

Recommended number of blood culture sets
Recommended number of blood culture sets

A contaminant will usually be present in only one bottle of a set of culture bottles, but if it’s a true bloodstream infection, multiple blood culture bottles/set will be positive.

Timing of Blood Collection

Timing of blood cultures is not considered a critical factor in determining bloodstream infections, as the diagnostic yield remains the same. Most of the researchers/authors concluded that “the overall volume of blood cultured was more critical in increasing organism yield than was timing”.

Guidelines recommend that the first two/three sets (2 bottles/set) of blood culture be obtained either at one time or over a brief time period (e.g. within 1 hour) from multiple venipuncture sites.

Collection and Dispensing of Blood for Culture

Blood for culture must be collected and dispensed aseptically with great care to avoid contaminating the specimen and culture medium.

  1. Using a pressure cuff, locate a suitable vein in the arm. Deflate the cuff while disinfecting the venipuncture site.
  2. 2 Wearing gloves, thoroughly disinfect the venipuncture site as follows:
    1. Using 70% ethanol, cleanse the skin over the venipuncture site in a circle approximately 5 cm in diameter, rubbing vigorously.  Allow air-drying.
    2. Starting in the center of the circle, apply a 2% tincture of iodine (or povidone-iodine) in ever-widening circles until the entire circle has been saturated with iodine. Allow the iodine to dry on the skin for at least 1 minute. The timing is critical, use a watch or a timer.
  3. Lift back the tape or remove the protective cover from the top of the culture bottle(s). Wipe the top of the bottle using an ethanol-ether swab.
    • Prior inspection of culture media bottles: Do not use a bottle of culture medium if it shows signs of contamination, i.e. broth appears turbid. Do not use a bottle of thioglycollate broth if it appears oxidized, i.e. more than a third of the top of the medium appears pink when the indicator in the medium is resazurin, or more than 20 mm down from the surface of the medium appears green-blue when the indicator is methylene blue. When oxidation has occurred, the medium must be reduced by steaming before it is used.
  4. Using a sterile syringe and needle, withdraw about 20 ml of blood from an adult* or about 2-5 ml from a young child. *Note: When an anaerobic culture is not indicated, collect 10 ml of blood.
  5. After removing the needle, cleanse the site with 70% ethanol again because many patients are sensitive to iodine.
  6. Insert the needle through the rubber liner of the bottle cap and dispense 10 ml of blood into the aerobic culture medium bottle.
    Do not change needles before injecting the blood into the culture bottle.
  7. When also culturing for anaerobes, dispense about 5 ml of blood into the thioglycollate culture medium containing 50 ml of broth.
  8. Using a fresh ethanol-ether swab, wipe the top of each culture bottle and replace the tape or protective cover(s). Without delay, mix the blood with the broth.
    Note: The blood must not be allowed to clot in the culture media because any bacteria will become trapped in the clot.
  9. Clearly label each bottle with the name and number of the patient, and the date and time of collection.
  10. As soon as possible, incubate the inoculated media. Protect the cultures from direct sunlight until they are incubated.

Tranport of Blood Culture Bottles

  • Do not refrigerate blood cultures. Generally hold at room temperature until processed, for a maximum of 4 h.
  • Refer to the manufacturer’s instructions for the appropriate method to store cultures prior to incubation in automated culture systems.

Rejection criteria

  1. Reject blood cultures that are received unlabeled.
  2. Do not process if the tube or bottle is cracked or broken.

References and further readings: 

  1. Blood Culture: A key Investigation for Diagnosis of Bloodstream Infections (BioMérieux)
  2. Bailey & Scott’s Diagnostic Microbiology, Forbes, 11th edition

Acharya Tankeshwar

Hello, thank you for visiting my blog. I am Tankeshwar Acharya. Blogging is my passion. As an asst. professor, I am teaching microbiology and immunology to medical and nursing students at PAHS, Nepal. I have been working as a microbiologist at Patan hospital for more than 10 years.

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