Nebraska Public Health Laboratory

1. Nebraska Public Health Laboratory 2008 CLSI M100-S18 update Paul D. Fey, Ph.D. Associate Professor/Associate Director Josh Rowland, M.T. (ASCP) State Training Coordinator

2. Agenda • Discuss 2008 M100-S18 major changes • Organism specific changes • KPC carbapenemase • Other AST issues and questions from discussion group

3. Issue 1-New Appendices • Appendix A -ESBL screening • Appendix B -Staphylococcus aureus susceptibility testing issues • Appendix C -Coagulase-negative Staphylococcus susceptibility testing issues • Appendix D -Screening for high-level Aminoglycoside resistance within the enterococci.

4. Tables 1 and 1A-Antibiotic reporting tables • Starting pages 24 (Disk diffusion) and 86 (MIC). Tables 1 and 1A. • Changes Table 1: • Tobramycin moved from group B (selective reporting) to group A (primary) in Enterobacteriaceae and Pseudomonas aeruginosa. • Ticarcillin moved from group A to group B in Enterobacteriaceae and Pseudomonas aeruginosa. • Macrolides, Clindamycin, Trimethoprim/sulfa. moved to group A under Staphylococcus. • Note that Daptomycin is MIC only-do not use disk diffusion disks • Acinetobacter spp. Ampicillin-sulbactam, ciprofloxacin, levofloxacin and gentamicin, tobramycin moved to group A.

5. Tables 1 and 1A-Antibiotic reporting tables • Starting pages 24 (Disk diffusion) and 86 (MIC). Tables 1 and 1A. • Changes table 1A • Note that “Streptococcus spp. other than Streptococcus pneumoniae” column has been split into two columns: 1) Streptococcus spp. Beta-hemolytic group (Groups A, B, C, and G) and 2) Streptococcus spp. Viridans group (Small colony forming b-hemolytic colonies [ S. anginosus] are included in this group).

6. Haze in zone of inhibition • New comments on M2-disk diffusion tables about appropriate methods to read zones of inhibition to certain antibiotics—i.e. Proteus mirabilis and swarming, trimethoprim-sulfamethoxazole testing, etc. Please read all comments if disk diffusion is common in your laboratory. • Page 52—”Organisms that show hazy growth throughout the zone of inhibition around the clindamycin disk should be reported as clindamycin resistant, whether or not they show a D-zone.” • To detect any type of colonies that may show resistance-Page 52-”When testing linezolid, disk diffusion zones should be examined using transmitted light. Organisms with non-susceptible results should be confirmed using an MIC method.”

7. Streptococcus pneumoniae • If you are performing disk diffusion to detect resistance in Streptococcus pneumoniae, read page 66. • “Penicillin MICs should be determined for those isolates with oxacillin zone diamters < 19 mm, because zones of < 19 mm occur with penicillin-resistant, intermediate, or certain susceptible strains based upon the meningitis and oral penicillin V interpretive criteria given in M7, Table 2G. Isolates should not be reported as penicillin-resistant or intermediate based solely on an oxacillin-zone < 19 mm.”

8. Streptococcus pneumoniae • Significant reporting changes regarding penicillin • Interpretive standards for parenteral penicillin non-meningitis, meningitis, and oral penicillin non-meningitis. • On meningitis isolates, only report meningitis interpretive standards. • On non-meningitis isolates (e.g. pneumonia or blood), report both meningitis and non-meningitis interpretive standards.

9. Ceftriaxone (meningitis)- <0.5 S Ceftriaxone (non-meningitis)- <0.5 S Erythromycin->1 R Levofloxacin- <0.5 S Penicillin (meningitis)-1 R Penicillin (non-meningitis)-1 S Penicillin oral (non-meningitis)-1 I Vancomycin-0.5 S Based on therapy category, different interpretive standards. Multiple comment suggestions in the CLSI These changes must reflected in your antibiogram. Streptococcus pneumoniae isolated from blood-penicillin MIC of 1

10. Staphylococcus aureus changes • Page 111. Cefoxitin MIC test to detect methicillin resistance—only reliable for S. aureus and S. lugdunensis. • “The results of cefoxitin MIC tests can be used to predict the presence of mecA-mediated resistance in S. aureus and S. lugdunensis. Isolates for which cefoxitin MICs are > 8 mg/ml should be reported as resistant. Isolates for which the cefoxitin MICs are < 4 mg/ml should be reported as oxacillin susceptible.”

11. Staphylococcus aureus changes • Page 114-Erythromycin-resistant and clidamycin-susceptible isolates. New approved way to detect inducible resistance to clindamycin. Well containing 4 mg/ml erythromycin and 0.5 mg/ml clindamycin—growth in the well indicates resistance to clindamycin.

12. KPC carbapenemase • Enzyme capable of hydrolyzing carbapenems (imipenem, ertapenem, meropenem, doripenem). • Isolated primarily on the east coast of US in ICUs. • Klebsiella pneumoniae and other Enterobacteriaceae. • KPC not easily detected using commercial susceptibility systems—some isolates have an MIC of 2 to the carbapenems (still susceptible) • KPC enzymes hydrolyze expanded-spectrum cephalosporins as well—therefore, all Enterobacteriaceae isolates that have an ESBL phenotype should be screened for a carbapenemase.

13. KPC carbapenemase • Ertapenem most useful carbapenem to detect KPC. • If ertapenem is not on your panel, suggest that all ESBL/AmpC-like Enterobacteriaceae (i.e. expanded-spectrum cephalosporin resistant) be tested with ertapenem (and other carbapenems) by disk diffusion or MIC method (E-test). • If isolate has MIC of > 2 mg/ml (susceptible isolates should have an MIC of < 0.5 mg/ml), contact physician and send isolate to reference laboratory. • Modified Hodge test • PCR for KPC