Microbiological examination of wounds

Sampling from open infected wounds

Sampling from open injuries requires a strict sequence of actions to avoid introducing normal skin microflora into the sample. First, the operative field is prepared: the skin strictly around the wound is treated with an antiseptic (70% ethanol is used), and the disinfectant is allowed to dry completely. Then, mechanical cleaning of the wound surface itself is initiated.

Using a sterile wipe, remove all surface detritus, accumulations of pus, and necrotic masses. Reliable material for microbiological examination can only be obtained from viable ("living") tissues.

A sterile swab is used for direct sampling. Using rotational circular movements, the swab is moved from the center of the lesion to its periphery. The optimal contact duration with the wound base is 5 to 10 seconds. It is critically important during instrument removal not to touch the surrounding intact tissues, mucous membranes, or skin edges under any circumstances. Immediately after taking the swab, it is immersed in a tube with transport medium to preserve microorganisms.

Specifics of working with deep wounds and abscesses

The algorithm for collecting samples from deep wounds and abscesses differs significantly. The preparatory phase involves careful removal of surface exudate (using a wipe moistened with saline or 70% alcohol), followed by disinfection and drying of the surface.

The preferred collection method in this case is syringe aspiration of contents from the very depth of the wound tract. If vesicles are present at the base of the defect, their liquid contents must be aspirated along with the base cells. In situations where free fluid is absent, sterile saline is injected into the wound cavity, followed by repeated aspiration of the resulting washings.

The obtained liquid aspirate is transferred to an empty sterile tube. Alternatively, a sterile swab may be thoroughly saturated with the collected exudate and then placed into the transport medium. When diagnosing established abscesses, additional samples are taken:

• Biopsy specimen: a piece of tissue from the pyogenic membrane (pushed deep into the transport medium using a swab).
• Microscopy: contents are applied to a glass slide with a separate swab for direct examination.

Transportation rules and primary inoculation

The success of bacteriological diagnostics largely depends on compliance with logistics. The optimal delivery time of the collected material to the microbiological laboratory is no more than 1 hour from the moment of collection. If immediate transportation is impossible, samples may be stored in a refrigerator, but the maximum delay must not exceed 2 hours.

Primary inoculation is performed in the laboratory. The material from the swab is distributed across a standard set of nutrient media:

1. 5% blood agar (for most pathogens);
2. Thioglycolate broth (specifically for cultivating anaerobes);
3. Sugar broth.

Not only swabs or liquid aspirates, but also solid tissue pieces delivered from the operating room are subject to mandatory inoculation.

Technique for inoculating tissue material

Solid samples (tissue biopsies, muscle fragments, skin areas, or bone sequestra) obtained during surgical debridement require a special approach. Initially, such tissues are placed in liquid media: thioglycolate broth (supporting the growth of both anaerobes and aerobes) and sugar broth, which serves as an enrichment medium.

To obtain isolated colonies of microorganisms even from complex microbial associations on solid media, the "swab-loop" technique is used. It consists of two stages:

• Stage 1 (Swab): First, a continuous "streak" is made across the entire diameter of the Petri dish using the swab containing the material. Then, the instrument is turned over, and a second "streak" is made with the other (clean) side, strictly parallel to the first, but in the opposite direction.
• Stage 2 (Loop): Using a bacteriological loop, the applied material is streaked out with frequent lines placed strictly perpendicular to the previously made "streaks".

Petri dishes with inocula are incubated under standard conditions: a temperature of 37 °C for 18–24 hours. If visual growth of microorganisms is absent after a day, the cultures are not discarded. They are left in the incubator for up to 5 days to avoid missing the growth of slow-growing bacteria.

Class and exam questions

What else to study on this topic

• Rationale for selecting media for primary inoculation (5% blood agar, thioglycolate broth, sugar broth).
• Specifics of sample collection in the presence of vesicles at the base of the defect.
• Detailed "swab-loop" inoculation technique for isolating single colonies.
• Algorithm for preparing the skin around the surgical site before taking a smear.
• Principles of microscopy of an individual smear in the diagnosis of formed abscesses.

Other topics in the «Clinical Microbiology» section

Prepared by WhiteCoat Pro faculty in accordance with the medical school curriculum. For educational purposes only.