Antimicrobial susceptibility testing using disc diffusion methods, A.

 

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Historical aspects and the concept of species-related interpretive zone diameter breakpoints

Antimicrobial susceptibility testing in the clinical laboratory is most often performed using the disc diffusion method. This method was originally standardized by Bauer et al. (the so called Kirby-Bauer method), and by Ericsson & Sherris.

A well-known authority on antimicrobial susceptibility testing, Dr Ronald Jones, USA, wrote: " ... the disk diffusion test continues to be the most versatile, broadly accurate, and reproducible AST test we can use in the clinical microbiology laboratory." [Jones, R.N. 1992. Clinical Microbiology Newsletter 14:33-37]

The Ericsson & Sherris method was adopted early as the method of choice for susceptibility testing in Sweden, but in those days the susceptibility categories were called 1, 2, 3, and 4. In 1978 this was changed by the Swedish reference group to the more international S, I, and R categories.

In 1976 we had started to plot zone diameter histograms species-wise for the different antibiotics in Lund. This revealed homogeneous populations of isolates representing wild type susceptible isolates as well as strains with various degrees of resistance. The new zone breakpoints from SRGA in 1978 did not always fit individual species so exceptions with special breakpoints were introduced locally in Lund. We concluded that a proper asignment of a susceptibility category of an isolate as S, susceptible, I, intermediate susceptible, or R, resistant, to a given antibiotic required knowledge about the distribution of zone diameters for that combination of bacterial species and antibiotic. The concept of species-related interpretive zone breakpoints for SIR categorization was introduced. The basic rule is that an interpretive zone breakpoint should never cut a homogeneous population of isolates in a zone histogram for a given combination of bacterial species and antibiotic, be it a fully susceptible population or one with decreased susceptibility.

Comparative quality control studies in the 1980:ies were performed by the Lund team involving all laboratories in Sweden. In two of the studies, Gunnar Kahlmeter, then a young doctor in the lab, was a team member. All these studies showed very clearly that there were interpretive problems using general breakpoints. The Swedish Reference Group for Antibiotics, SRGA, then appointed Gunnar Kahlmeter in 1986 to chair a methodology subgroup to further analyze the experiences gained in Lund and to produce experimental material from several other laboratories to test the concept of species-related interpretive breakpoints, a concept which was later adopted as a national standard in Sweden. For full details of the SRGA standard, interpretive breakpoints, populations of isolates with both MIC and zone diameter distributions, and other valuable information FREE OF CHARGE, both in English and in Swedish, see the SRGA home page.

Species-specific interpretation was not new, but had earlier only been suggested for occasional combinations of species and antibiotics, e.g. carbenicillin and P.aeruginosa in 1974 and chloramphenicol and P.mirabilis in 1980. Our extensive histogram studies of the years showed the need for a more general concept of species-related interpretive breakpoints. It should also be mentioned that Thomas F. O'Brien had pioneered studies of zone diameter histograms already in the 1960:ies.

All histogram studies so far had used simple visual inspection with comparison of interpretive breakpoints to evaluate possible problems. We felt the need for a more objective method to analyze histograms species-wise and then came up with "single-strain regression analysis" to produce species-specific regression lines,
(see next page).

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Page updated by GK, 2015-10-09