About the CF Referral Center

Introduction
The CF Referral Center for Susceptibility & Synergy Studies, or better known as "The Synergy Lab," was started in 1991 as a research project funded by the CF Foundation intended to assist CF clinicians treating CF patients infected with multidrug-resistant organisms. Thus, we perform standardized susceptibility and synergy studies for multidrug-resistant Pseudomonas isolated from patients with CF. These are in vitro studies of antibiotic activity and have not been correlated with clinical efficacy at this time. Results from these studies should be used at the discretion of the treating physician.
The CF Referral Center's microbiology laboratory and its procedures are compliant with CLIA‡ standards. In addition, our medical informatics and information systems are compliant with HIPAA/X12N guidelines.‡‡ Copies of certifications are available upon request. Patients are not charged for this service as The Synergy Lab is funded by the CF Foundation.

Overview of the CF Referral Center
As of September 2006, the CF Referral Center has studied >30,000 gram negative isolates from CF patients from 175 CF centers in the United States. Frozen stock of these isolates is available for other investigators.

Processing and Reporting of Studies
Specimens are received daily and processed within 24 hours of receipt Monday through Friday. Detailed instructions for sending isolates can be found at: "How to Submit Specimens". The turn-around-time for preliminary results is 48-72 hours after isolate receipt. These results are available online via our secure 3WCFSDC server for registered end-users to download.

Susceptiblity Studies
Susceptibility testing is performed by microbroth dilution using frozen commercially prepared microtiter plates (Microtech Medical Systems, Inc., Aurora, CO). This assay has been endorsed by the Clinical and Laboratory Standards Institute (CLSI)† to determine the antimicrobial susceptibility of multidrug-resistant P. aeruginosa isolated from patients with CF (Saiman et.al, 1999, J Clin Microbiol). However, there can be discrepancies between the antibiograms obtained by the referring laboratories and the Synergy laboratory if different susceptibility testing methods are used (Burns JL, et.al, 2000, J Clin Microbiol; Burns JL, et.al, 2001, Diagn Microbiol Infect Dis).

The minimal inhibitory concentration (MIC) is recorded and the MIC interpretation is the CLSI interpretive standard. A list of antimicrobial agents studied for Pseudomonas spp. is shown below. High concentrations of 256 µg/ml of tobramycin are also tested as such concentrations may be achieved by aerosolized antibiotic administration (Ramsey et al, N Eng J Med 1995).

1-Ticarcillin+Clavulanate; 2-Piperacillin+Tazobactam

Synergy Studies
Checkerboard synergy studies are performed with checkerboard dilutions of antibiotic pairs in tested at clinically achievable concentrations of drugs using frozen commercially prepared microtiter plates (Microtech Medical Systems, Inc., Aurora, CO). The same inoculum is used for the susceptibility studies and synergy studies.

Susceptibility and synergy panels are incubated at 35°C for 18 hours and slowly growing strains are incubated for an additional 18-24 hours. Following incubation, the plates are placed on a mirrored surface and turbidity is visualized using the microtiter test reading mirror (DYNEX Technologies, Chantilly, VA).

The fractional inhibitory concentration (FIC) for the synergy studies is calculated by comparing the activity of an agent in combination (MIC of agent A + agent B) with the activity of the antibiotic alone (MIC of agent A or agent B):

FICs are interpreted as follows:

Synergy combinations tested for P. aeruginosa.:

Additional Activities of the CF Referral Center

• We have IRB approval to supply investigators from academic institutions, biotechnology companies, and pharmaceutical companies with technical and clinical expertise and with de-identified clinical isolates.
• We can assist with study design, perform in vitro susceptibility and synergy studies with new agents, and perform data analysis and manuscript submission (Chen et.al, 2005, Antimicrob Agents Chemother).
• We assist in the monitoring of national standards for microbiological processing of respiratory tract specimens from CF patients by interfacing with regulatory agencies and / or collaborating with investigators from academic institutions.
• We study trends in antibiotic susceptibility of pathogens in CF patients.

Recent Regulatory Changes by CLSI
Please note that the CLSI has recently recommended a limited number of agents for susceptibility testing by local clinical microbiology laboratories (see below, Clinical and Laboratory Standards Institute. 2006. Performance standards for antimicrobial susceptibility testing; 16th informational supplement, vol. 26. Clinical and Laboratory Standards Institute, Wayne, PA.).

While CF strains are often more resistant than non-CF strains, these recommendations are nevertheless applicable to isolates from patients with CF.

In addition, we have recently reviewed the aggregate results of our synergy tests. Numerous combinations demonstrated synergistic activity against S. maltophilia and most of these agents are recommended by the CLSI (San Gabriel et.al, 2004, Antimicrob Agents Chemother). However, fewer synergistic combinations for B. cepacia complex are noted (Zhou et.al, 2006, Antimicrob Agents Chemother, In press). Thus, it is likely that checkerboard synergy studies have limited utility against B. cepacia complex strains.

Other Considerations
The Director of the CF Referral Center, Dr. Lisa Saiman, continues to provide clinical advice to CF centers and individual clinicians. She also serves as a consultant to several organizations on CF care-related issues.

Antibiotics are added or deleted as new agents or studies become available. For details on the current testing methods by the CF Referral Center, or if you need further assistance please call our laboratory at (212) 305-1991 and/or email us at synergy@columbia.edu.

Please let us know if you have any questions, comments or suggestions.

Thank you in advance for using the CF Referral Center. We hope that our services will continue to be useful.



Lisa Saiman, MD, MPH

†Clinical and Laboratory Standards Institute, Wayne, PA . ‡Clinical Laboratory Improvement Act of 1988 by the Centers for Medicare & Medicaid Services.
‡‡The Health Insurance Portability and Accountability Act of 1996 (HIPAA). Zosyn® is a registered trademark of Lederle Laboratories. Timentin® is a registered trademark of SmithKline Beecham Pharmaceuticals. Bactrim® is a registered trademark of Hoffmann-LaRoche, Inc. CFF® is a registered trademark of the Cystic Fibrosis Foundation.
Testing procedures are subject to change as new agents or studies become available.