Antimicrobial Susceptibility Testing of Aquatic Bacteria: Quality Control Disk Diffusion Ranges for Escherichia coli ATCC 25922 and Aeromonas salmonicida subsp. salmonicida ATCC 33658 at 22 and 28°C

Abstract

Quality control (QC) ranges for disk diffusion susceptibility testing of aquatic bacterial isolates were proposed as a result of a multilaboratory study conducted according to procedures established by the National Committee for Clinical Laboratory Standards (NCCLS). Ranges were proposed for Escherichia coli ATCC 25922 and Aeromonas salmonicida subsp. salmonicida ATCC 33658 at 22 and 28°C for nine different antimicrobial agents (ampicillin, enrofloxacin, erythromycin, florfenicol, gentamicin, oxolinic acid, oxytetracycline, ormetoprim-sulfadimethoxine, and trimethoprim-sulfamethoxazole). All tests were conducted on standard Mueller-Hinton agar. With >95% of all data points fitting within the proposed QC ranges, the results from this study comply with NCCLS guidelines and have been accepted by the NCCLS Subcommittee for Veterinary Antimicrobial Susceptibility Testing. These QC guidelines will permit greater accuracy in interpreting results and, for the first time, the ability to reliably compare susceptibility test data between aquatic animal disease diagnostic laboratories.

Quality control ranges for testing broth microdilution susceptibility of Flavobacterium columnare and F. psychrophilum to nine antimicrobials

“…This newly standardized AST method for Flavobacteria and the MIC QC ranges provided in Table 6 will be included in the next edition of the CLSI M49-A guideline. We hope that our work as well as the work of Miller et al (2003Miller et al ( , 2005 will serve as helpful resources to aid in continued development of more standardized AST methods for other important aquatic bacterial pathogens. …”

“…Proper QC procedures with expected test results are required to confirm method performance. Miller et al (2003Miller et al ( , 2005 established the first standard reference disk diffusion and MIC testing methods for non-fastidious aquatic bacterial pathogens (CLSI 2006a,b). These organisms include members of Enterobacteriaceae and Vibrionaceae, and Aeromonas sal monicida, A. hydrophila, and other mesophilic aero monads, Pseudomonas spp., Plesiomonas shi gelloides, and Shewanella spp.…”

“…They represent an important group that need standard anti microbial susceptibility testing (AST) methods. The Clinical and Laboratory Standards Institute (CLSI) provides 2 consensus-approved guidelines with reference methods and quality control (QC) criteria for disk diffusion and broth dilution susceptibility testing of non-fastidious aquatic bacteria (CLSI 2006 a,b) based on the work of Miller et al (2003Miller et al ( , 2005. CLSI groups fastidious aquatic bacterial pathogens that have similar growth condition requirements, and offers potential media and incubation modifications; however, no standard reference methods or QC criteria have been developed to date.…”

Impact of inter-lab variation on the estimation of epidemiological cut-off values for disc diffusion susceptibility test data for Aeromonas salmonicida

“…However, increasing standardisation has been demonstrated only to minimise but not eliminate inter-laboratory variance (Kronvall, 2003). Inter-laboratory studies have been published of the data generated by three disc diffusion test protocols designed to examine bacteria associated with aquatic animals (Miller et al, 2003;NicGabhainn et al, 2003;Huys et al, 2005). All have reported significant inter-laboratory variance between the data generated by laboratories using the same test protocols.…”

“…All have reported significant inter-laboratory variance between the data generated by laboratories using the same test protocols. The width of the acceptable ranges for control strains in M42-A (CLSI, 2006a), which were generated from the standard deviations of data generated in nine laboratories (Miller et al 2003), illustrate the extent to which inter-laboratory variation remains a problem even after rigorous standardisation. For A. salmonicida ACTC 33658 incubated at 22 °C for 44-48 h the width of these acceptable ranges vary from 11-16 mm (median 13 mm).…”