Effect of Neutralizing Culture Media on Positive Colonization Determinations for Antimicrobial Catheters

Background: Catheter related blood stream infections (CRBSIs) cause significant mortality and increase the cost of healthcare.  Two antimicrobial CVCs have CDC (Centers for Disease Control and Prevention) category 1B recommendations for prevention of CRBSIs based on clinical trial evidence.  The CRBSI definition from CDC (2002 guidelines; Appendix A) requires clinical infection symptoms, a positive blood culture from a peripheral vessel, and a positive determination of catheter colonization by a matching organism.  The most stringent method for positive catheter colonization determination is quantitative catheter segment culture in excess of 1000 cfu/ml.  Some clinical trials on CVCs treated with potent antibiotics have not used neutralizing culture media when culturing explanted catheter segments. This creates the possibility that antimicrobial carryover from the catheter to the culture broth or agar can falsely lower colonization counts due to latent inhibition of recovered organisms.  Incorrect attribution of infections to CVCs due to erroneous negative colonization determinations can lead to under-reporting of CRBSI rates from clinical trials.    

Objective: This study assessed whether excluding neutralizing media in antibiotic CVC culturing protocols could give erroneous negative results for catheter colonization due to antimicrobial carryover.

Methods: Minocycline (M) and Rifampin (R) treated (triplicate) catheter segments were exposed to 5x10⁶ cfu/ml of Staphylococcus epidermidis (SE) ATCC #35983 in trypticase soy (TS) broth for 24 hrs.   Quantitative culture and plating was performed following the challenge by first sonicating the catheter in broth followed by dilution, plating on agar, incubation (24 hours), and colony counting.   The effect, on quantitative recovery, of sonicating in neutralizing Dey-Engle (DE) broth and non-neutralizing TS broth was measured.  In addition, the effect of plating onto DE or TS agar was also determined.   Minimum Inhibitory Concentrations (MICs) of M and R in TS and DE broths were measured by serial dilution to determine DE neutralization capacity. 

Results: :  MICs of M and R for SE in TS broth were below 0.06 μg/ml.   In DE broth MICs increased to above 1 μg/ml.  No colonies were recovered following sonicating in either DE or TS broths and plating onto TS agar.  The mean recovered colony counts following sonicating in either DE or TS both and plating onto DE agar was greater than 7000 cfu/ml. 

Conclusions: Excluding neutralizing agents in culture media can lead to false negative colonization determinations for antimicrobial CVCs based on the CDC limit of 1000 cfu/ml for a positive finding.    For highly potent antibiotics, use of a neutralizing agar during plating is needed to minimize antimicrobial carryover if the neutralizing capacity of the sonication broth may be exceeded.