Wound Culture

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Wound Culture

Definition

A wound culture is a diagnostic laboratory test in which microorganisms from an infected wound are grown in the laboratory on media and identified. Wound cultures always include aerobic culture, but direct smear evaluation (Gram stain ) and anaerobic culture are not performed on every wound. These tests are performed when indicated or requested by the physician.

Purpose

The purpose of a wound culture is to isolate and identify microorganisms causing an infection of the wound, and to identify antibiotics that will be effective in destroying the organism.

Preparation

A biopsy sample is usually preferred by clinicians, but this is a moderately invasive procedure and may not always be feasible. The patient is prepped by cleansing the area with a sterile solution such as saline. Antiseptics such as ethyl alcohol are not recommended since they will kill bacteria and results will be negative. The patient is given a local anesthetic and the tissue is removed using a cutting sheath. Pressure is applied to the wound to control bleeding. Needle aspiration is less invasive and is a good technique to use in wounds where there is little loss of skin such as puncture wounds. Skin around the wound is cleaned with an antiseptic to kill bacteria on the skin's surface and a small 22 gauge needle is inserted. The clinician should pull back on the plunger and then change the angle of the needle two or three times to remove fluid from different areas of the wound. This procedure may be painful for the patient, so many initial cultures are done with the swab technique. The nurse should clean the wound area with sterile saline and moisten a sterile swab with sterile saline. The tip is inserted into the wound and rotated with pressure applied. The pressure will give a better yield of the fluid that is deeper in the wound. The swab used for anaerobic culture should be oxygen-free. After all three procedures, the wound should be cleaned thoroughly and bandaged.

Tissue specimens collected by biopsy should be placed in a screw capped vial containing a small amount of sterile saline to keep the tissue moist. The anaerobic sample should be placed in a gassed out vial that may contain prereduced medium or a gassed out bag and sealed. Syringes should be tightly capped immediately following aspiration. A common practice for anaerobic culture is to inoculate an anaerobic blood culture bottle at the point of care to insure the sample is not exposed to air. Several swabs (at least three) should be collected. One swab is placed in Stuart, Cary-Blair, or Amies transport medium for aerobic culture and another in PRAS transport medium for anaerobic culture. One swab is placed in a clean dry envelope or tube for direct smear examination.

Description

Wounds are injuries to body tissues caused by physical trauma or disease processes including surgery, diabetes, burns, punctures, gunshots, lacerations, bites, bed sores and broken bones. Types of wounds include:

Abraded: Caused by abrasion such as falling on concrete.
Contused: A bruise or contusion.
Incised: Caused by a clean cut, as by a sharp instrument.
Lacerated: Caused by a laceration, tearing of the skin or tissues.
Nonpenetrating: Injury caused without disruption of the surface of the body. These wounds are usually in the thorax or abdomen and can also be termed blunt trauma wounds.
Open: A wound in which tissues are exposed to the air.
Penetrating: Disruption of the body surface and extension into the underlying tissue.
Perforating: A wound with an exit and an entry, such as a gunshot wound.
Puncture: A wound formed when something goes through the skin and into the body tissues. This wound has a very small opening but can be very deep.

The chance of a wound becoming infected is dependent upon the nature, size, and depth of the wound; its proximity to and involvement of nonsterile areas such as the skin and gastrointestinal tract; the opportunity for organisms from the environment to enter the wound; and the immunological and general health status of the person. Skin and body compartmentalization prevent many infections. In general, acute wounds are more prone to infection than chronic wounds. Wounds with a large loss of body surface such as abrasions are also easily infected. Puncture wounds may permit the growth of micro organisms since there is a break in the skin with minimal bleeding and they are hard to clean. Deep wounds, closed-off from oxygen, are an ideal environment for an anaerobic infection to develop. Foul-smelling odor, gas, or gangrene at the infection site are signs of an infection caused by an anaerobic bacteria. Surgical wounds can cause infection by introducing bacteria from one body compartment into another.

Diagnosing infection in a wound may be difficult. One of the cardinal signs the clinician looks for is slow healing. Within hours of injury, most wounds will display a release of fluid called an exudate. This fluid contains compounds that aid in healing and is normal. It should not be present 48-72 hours after injury. Exudate indicative of infection may be thicker than the initial exudate and may also be purulent (containing pus) and foul smelling. Clinicians will look at color, consistency and the amount of exudate to monitor early infection. In addition, infected wounds may display skin discoloration, swelling, warmth to touch and an increase in pain.

Wound infection prevents healing, and the microorganisms can spread from wounds to other body parts, including the blood. Infection in the blood is termed septicemia and can be fatal. Symptoms of a systemic infection include a fever and rise in white blood cells, along with confusion and mental status changes in the elderly. It is important to treat the infected wound early with a regimen of antibiotics to prevent further complications.

Wound infections often contain multiple organisms including both aerobic and anaerobic grampositive cocci and gram-negative bacilli and yeast. The most common pathogens isolated from wounds are Streptococcus group A, Staphylococcus aureus, Escherichia coli, Proteus, Klebsiella, Pseudomonas, Enterobacter, Enterococci, Bacterioides, Clostridium, Candida, Peptostreptococcus, Fusobacterium, and Aeromonas.

A Gram stain is prepared by rolling the smear across the center of a glass slide or dropping a liquid specimen onto the center and allowing it to air dry. The initial Gram stain is used to evaluate the adequacy of the specimen; estimate the amount of any bacteria, yeast, or fungus present; and determine whether a specialized culture medium is required based upon the appearance of the organisms found. For example, the Gram stain may reveal gram positive filamentous bacteria suggestive of Norcardia that requires special growth medium.

The tissue used for the tests is obtained by three different methods, tissue biopsy, needle aspiration or the swab technique. The biopsy method involves the removal of tissue from the wound using a cutting sheath. The piece of tissue is transported to the laboratory where it must be liquified. This is done by adding approximately 1 mL of liquid medium and grinding the tissue in a blender or grinder until it forms a thick homogenized liquid. This is vortexed and dispensed onto solid media and into broth with a sterile pipet. Samples aspirated by syringe can be injected directly into broth and dispensed onto solid media. The swab technique is most commonly used but contains the least amount of specimen, and therefore, recovery is lower than with biopsied tissues or aspirates. The swab is pressed against the transport tube and the suspen sion of transport media is transferred to broth and solid media with a sterile pipet.

Wound specimens are cultured on both nonselective enriched and selective media. Cultures for anaerobes should include anaerobic sheep blood agar supplemented with vitamin K and hemin for general isolation; kanomycin-vancomycin laked blood agar for Bacteroides spp.; phenylethyl alcohol (PEA) or colistin-nalidixic acid (CNA) anaerobic sheep blood agar to suppress gram-negative bacilli; and thioglycolate broth with hemin and vitamin K for slow growing organisms, especially if tissues or aspirates are being cultured. Anaerobic media are inoculated inside a glove box or in an anaerobic (degassed) holding jar and incubated at 96.8 °F (36 °C) in the absence of oxygen for five to seven days. Aerobic culture should include inoculation of sheep blood agar for general growth; chocolate agar for isolation of Haemophilus; MacConkey agar for isolation of enteric gram negative bacilli; CNA or PEA blood agar for gram-positive cocci; and potato dextrose agar with antibiotics for isolation of yeast. Cultures are incubated in humid air at 96.8 °F (36 °C) for 48 hours (except for chocolate agar which is incubated in 5-10% carbon dioxide). Cultures are examined each day for growth and any colonies are Gram stained and subcultured (i.e., transferred) to appro priate media. The subcultured isolates are tested via appropriate biochemical identification panels to identify the species present. Organisms are also tested for antibiotic susceptibility by the microtube broth dilution or Kirby Bauer method. The selection of antibiotics for testing depends upon the organism isolated (i.e., gram-negative versus gram-positive, aerobe versus anaerobe).

Results

The initial Gram stain result is available the same day, or in less than an hour if requested by the physician. An early report, known as a preliminary report, is usually available after one day. After that, preliminary reports will be posted whenever an organism is identified. Cultures showing no growth are signed out after two to three days unless a slow growing mycobacterium or fungus is found. These organisms take several weeks to grow and are held for four to six weeks. The final report includes complete identification, an estimate of the quantity of the microorganisms, and a list of the antibiotics to which each organism is sensitive and resistant.

Complications

The physician may choose to start the person on an antibiotic before the specimen is collected for culture. This may alter results, since antibiotics in the person's system may prevent microorganisms present in the wound from growing in culture. In some cases, the patient may begin antibiotic treatment after the specimen is collected based upon Gram stain results or clinical findings. The antibiotic chosen may or may not be appropriate for one or more organisms recovered by culture.

KEY TERMS

Aerobe— Bacteria that require oxygen to live.

Agar— A gelatinous material extracted from red algae that is not digested by bacteria. It is used as a support for growth in plates.

Anaerobe— Bacteria that live only where there is no oxygen.

Antibiotic— A medicine that can be used topically or taken orally, intramuscularly or intravenously to limit the growth of bacteria.

Antimicrobial— A compound that prevents the growth of microbes which may include bacteria, fungi and viruses.

Antimycotic— A medicine that can be used to kill yeast and fungus.

Antiseptic— A compound that kills all bacteria, also known as a bactericide.

Broth— A growth mixture for bacteria. Different compounds such as sugars or amino acids may be added to increase the growth of certain organisms. Also known as media.

Exudate— Any fluid that has been released by tissue or its capillaries due to injury or inflammation.

Gram stain— A staining technique used in micro biology to identify and classify bacteria. The organisms will stay purple if gram-positive or counterstain pink if gram-negative. The Gram stain result depends upon the chemical composition of the bacterial cell wall.

Normal flora— The mixture of bacteria normally found at specific body sites.

Purulent— Contianing, consisting of or forming pus.

Pus— A fluid that is the product of inflammation and infection containing white blood cells and debris of dead cells and tissue.

Nurses must be very careful when finishing a wound culture collection to make sure the wound has been cleaned thoroughly and is bandaged properly. It is important to watch for bleeding and further infection from the procedure. In addition, patients may be in pain from the manipulation so pain killing drugs such as acetaminophen may be advised.

Health care team roles

Wound culture requires the expertise of many clinicians including nurses, doctors and microbiologists. A physician requests the wound culture and is responsible for specimen collection and antibiotic selection. The physician may be assisted by a nurse, nurse practitioner, or physician assistant. Nurses should inform the patient about the testing procedure and what pain to expect. They should clean the wound thoroughly afterwards, bandage it correctly and watch for signs and symptoms of further infection. Doctors should monitor the patient closely for signs of systemic infection and be prepared to repeat the procedure if the patient does not respond to a course of antibiotics. Cultures are performed by clinical laboratory scientists/medical technologists who specialize in clinical microbiology.