FAS ENDOLUMINAL BRUSH
RECOMMENDED MICROBIOLOGY TECHNIQUE:
Assessing Catheter Lumen Biofilm for the Presence of Microbial Growth or Colonisation
Introduction:
One of the major attributes of the FAS Endoluminal Brush product line is its ability to obtain a biofilm sample from internal lumens of Central Venous Catheters (CVC) while the catheters remain in-situ. The Endoluminal Brush is currently the only tool which enables clinicians to directly assess the microbial status of a lumen without removing the catheter from a patient – which ultimately can prevent catheters from being removed and replaced unnecessarily.
Standard microbiology methods (plate culture) may be used to test the biofilm sample retrieved by an Endoluminal Brush. Although the Endoluminal Brush is intended to be used while catheters are in-situ, it may also be used be used in an in-vitro or laboratory situation to assess catheters which have already been removed from patients. In this case the Endoluminal Brush technique is superior to other techniques, such as the Cleri Method, which are designed to examine intralumenal bacteria since there is no risk of contamination from the external surface.
PLATE CULTURE METHOD
Purpose
Isolates and quantitates microorgansims (colony forming units) which may be identified by using routine microbiology laboratory methods.
Materials Required
Ø Brush with Biofilm Sample in test tube or suitable container
Ø Sterile Phosphate Buffered Saline (PBS)
Ø 2 Blood Agar Plates
Ø 1 Sabouraud Plate
Ø Sterile Plastic Spreaders or equivalent
Ø 10 and 100 ul micropipettes and tips
Ø Waterbath Sonicator and / or Vortex Mixer
Ø Incubator
Preliminary Instructions
1. Retrieve Biofilm sample by following the FAS Endoluminal Brush Instructions for Use.
2. Place the Brush / Biofilm sample into the test tube provided with the kit or an equivalent container.
Procedure
1) Add 1 ml of Sterile PBS to the container holding the Brush / Biofilm sample (“Sample”).
2) Sonicate the Sample in a sonicating waterbath for 1 minute (Room Temperature).
Note: If a sonicating waterbath is not available, proceed directly to Step 3.
3) Vortex for 15 seconds.
Note: If a sonicating waterbath was not available, vortex for 1 minute.
4) Transfer 10 ul of the PBS / Biofilm solution on to Blood Agar Plate and another 10 ul on to a Sabouraud Plate. Spread over the surface of the plate using a sterile plastic spreader.
5) Repeat Step 4 using 100 ul of PBS / Biofilm solution on to the second Blood Agar Plate.
Note: Optional: Retain the residual PBS / Biofilm solution if performing the Cytocentrifuge Staining Technique.
6) Incubate all plates overnight at 37o C.
7) Examine plates for bacterial / yeast growth, record and report growth to clinicians according to the hospital’s standard procedures.
8) Reincubate and re-examine plates at 48 hours. Record and report results as above.
CAUTION: If a Biofilm sample has been taken from a catheter which is impregnated internally with antimicrobial compounds, then special procedures are necessary so that microorganisms are not inhibited during the culture procedure – leading to erroneous results. It is recommended that plates be made with culture media containing neutralising agents that prevent inhibition. A suitable media has been defined by Schmitt et al (1996) J Clin Microbiol 34 508-511.
Interpretation of Quantitative Culture: Data collected and evaluated has provided the following guidelines for the interpretation of culture results (Reference: Kite: J Clin Path 1997. 50:278-281):
< 100 CFU: Not clinically significant
> 100 CFU: Clinically significant
> 100,000 CFU: Diagnostic of CRS
