Multisite Reproducibility of the Broth Microdilution Method for Susceptibility Testing of Nocardia Species

Abstract

Antimicrobial susceptibility testing (AST) of clinical isolates of Nocardia is recommended to detect resistance to commonly used antimicrobial agents; such testing is complicated by difficulties in inoculum preparation and test interpretation. In this study, six laboratories performed repetitive broth microdilution testing on single strains of Nocardia brasiliensis, Nocardia cyriacigeorgica, Nocardia farcinica, Nocardia nova, and Nocardia wallacei. For each isolate, a total of 30 microdilution panels from three different lots were tested at most sites. The goal of the study was to determine the inter-and intralaboratory reproducibility of susceptibility testing of this group of isolates. Acceptable agreement (>90% agreement at ؎1 dilution of the MIC mode) was found for amikacin, ciprofloxacin, clarithromycin, and moxifloxacin. After eliminating MIC values from single laboratories whose results showed the greatest deviation from those of the remaining laboratories, acceptable agreement was also found for amoxicillin-clavulanic acid, linezolid, minocycline, and tobramycin. Results showed unsatisfactory reproducibility of broth microdilution testing of ceftriaxone with N. cyriacigeorgica and N. wallacei, tigecycline with N. brasiliensis and N. cyriacigeorgica, and sulfonamides with N. farcinica and N. wallacei. N. nova ATCC BAA-2227 is proposed as a quality control organism for AST of Nocardia sp., and the use of a disk diffusion test for sulfisoxazole is proposed as a check of the adequacy of the inoculum and to confirm sulfonamide MIC results. N ocardia species are ubiquitous in the environment and have been implicated in a variety of human infections, especially in immunocompromised patients. In these patients, infection most frequently begins in the lungs, but can quickly disseminate to nearly every organ of the body. Nocardia can also cause disease in the immunocompetent host, in whom it is usually introduced through traumatic injury (2).Nocardia species are known to vary in their susceptibilities to various antimicrobial agents, and some species, especially Nocardia farcinica and members of the Nocardia transvalensis complex, have been shown to be particularly resistant to commonly used antimicrobial agents. Therefore, susceptibility testing should be performed on any Nocardia isolate considered to be of possible clinical significance.In 2003, the Clinical and Laboratory Standards Institute (CLSI) published an approved standard (M24A) for susceptibility testing of mycobacteria, nocardiae, and other aerobic actinomycetes, which specifies the broth microdilution method as the recommended procedure for susceptibility testing of aerobic actinomycetes (6). This document has recently been updated (5). Several years of experience in testing Nocardia isolates at the University of Texas Health Science Center and at the National Institutes of Health have demonstrated difficulties that may occur in inoculum preparation due to the propensity for clumping demonstrated by these organisms. In addition, determination of susc...

Antimicrobial Susceptibility among Clinical Nocardia Species Identified by Multilocus Sequence Analysis

“…While in-house prepared panels are specified in that document, we have demonstrated the suitability of the Sensititre Rapmyco microdilution panel for the determination of antimicrobial susceptibility patterns of clinical isolates of Nocardia spp. MIC values for the three control strains, N. nova ATCC BAA-2227, N. asteroides ATCC 19247 T , and N. farcinica ATCC 23826, were highly reproducible and correlated well with expected resistance profiles (7,20,21). The MIC values are published here to assist other clinical laboratories in implementing quality control guidelines for the validation of Nocardia antimicrobial susceptibility testing assays using this commercial panel.…”

“…Prior to this study, non-Nocardia, rapidly growing bacteria were recommended as QC strains for susceptibility testing, despite their very different growth characteristics during susceptibility testing. As an outcome of the work of Conville et al (7), N. nova ATCC BAA-2227 was recommended as the Nocardia reference strain for antimicrobial susceptibility testing of Nocardia spp. Our testing of N. nova ATCC BAA-2227 showed that not only were our results reproducible over 20 repeat tests but we also obtained MICs that were within the mode Ϯ 1 dilution for all antimicrobials tested in the Conville et al study (7), except for two drugs (amikacin and imipenem) where the Sensititre Rapmyco panel did not include dilutions as low as those on the in-house study panel.…”

“…Previously identified species have been reclassified as species complexes encompassing multiple species, and numerous novel Nocardia species have been identified (5). Currently, 87 species are enumerated in the List of Prokaryotic Names with Standing in Nomenclature (LPSN) (http://www.bacterio.net/nocardia.html), many of which are clinically significant (1).However, data on antimicrobial susceptibility has lagged behind the advances in taxonomy, with only a few reports providing recent data on newer antimicrobials (4,6,7,8,9,10,11,12,13). These reports largely focus on the species traditionally associated with clinical infections, such that antimicrobial susceptibility data are not available for a large number of newly identified, but clinically relevant, Nocardia species that are isolated less frequently in the clinical laboratory.…”

Sulfonamide Resistance in Isolates of Nocardia spp. from a U.S. Multicenter Survey

“…In a recent CLSI multicenter study among six laboratories using the same strains of Nocardia and the same lot numbers of susceptibility panels, media, etc., multiple laboratories reported discrepancies in MIC interpretations for sulfonamides (5). This multicenter study showed that careful training and close scrutiny of the 80% inhibition endpoint were necessary to produce accurate MIC results.…”

“…The current study and the CLSI multicenter study emphasized the necessity for laboratory proficiency testing with unusual organisms, such as Nocardia (5). Unfortunately, there is no standardized proficiency testing program currently available, including from the College of American Pathologists (CAP).…”