Sample Collection from Open Infected Wounds
Collecting samples from open injuries requires a strict sequence of steps to prevent contamination with normal skin flora. First, the surgical field is prepared: the skin strictly around the wound is treated with an antiseptic (70% ethyl alcohol is used), and the disinfectant must be allowed to dry completely. Next, mechanical cleaning of the wound surface is performed.
Using a sterile gauze pad, all surface debris, pus accumulations, and necrotic masses must be removed. Reliable material for microbiological examination can only be obtained from viable ("living") tissue.
A sterile swab is used for direct collection. Rotating circular movements are made 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 when withdrawing the instrument never to touch surrounding intact tissue, mucous membranes, or skin edges. Immediately after sampling, the swab is placed into a transport medium tube to preserve the microorganisms.
Working with Deep Wounds and Abscesses
The algorithm for sampling deep wounds and abscesses is significantly different. The preparation stage involves gently removing surface exudate (using a gauze pad moistened with normal saline or 70% alcohol), followed by disinfecting and drying the surface.
The preferred collection method in this case is syringe aspiration of contents from the deepest part of the wound tract. If vesicles are present at the base of the defect, their fluid contents must be aspirated along with the cells of the base. In situations where free fluid is absent, sterile normal saline is injected into the wound cavity, followed by repeated aspiration of the resulting wash.
The obtained liquid aspirate is transferred to an empty sterile tube. Alternatively, a sterile swab may be heavily saturated with the collected exudate and then placed into a transport medium. When diagnosing established abscesses, additional samples are taken:
- Biopsy: A piece of tissue from the abscess wall (pushed deep into the transport medium using a swab).
- Microscopy: Using a separate swab, the contents are applied to a glass slide for direct examination.
Transport Rules and Primary Inoculation
The success of bacteriological diagnostics largely depends on adhering to logistics. The optimal delivery time for collected material to the microbiology laboratory is no more than 1 hour from the moment of collection. If immediate transport 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. Material from the swab is distributed onto a standard set of nutrient media:
- 5% sheep blood agar (for most pathogens);
- Thioglycolate broth (specifically for culturing anaerobes);
- Nutrient broth.
Mandatory inoculation applies not only to swabs or liquid aspirates, but also to solid tissue fragments delivered from the operating room.
Technique for Tissue Inoculation
Solid specimens (tissue biopsies, muscle fragments, skin sections, or bone sequestra) obtained during surgical debridement require a special approach. Initially, such tissues are placed in liquid media: thioglycolate (supporting the growth of both anaerobes and aerobes) and nutrient broth, which serves as an enrichment medium.
To obtain isolated microbial colonies even from complex microbial associations on solid media, the "swab-loop" technique is used. It consists of two stages:
- Stage 1 (Swab): First, the swab containing the material is drawn in a continuous streak across the entire diameter of the Petri dish. Then the instrument is flipped, and the other (clean) side is used to make a second streak, strictly parallel to the first, but in the opposite direction.
- Stage 2 (Loop): Using a bacteriological loop, the applied material is distributed in frequent strokes oriented 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 microbial growth is absent after 24 hours, the cultures are not discarded. They are kept in the incubator for up to 5 days to avoid missing slow-growing bacteria.