Influence of disc content on zone sizes obtained in standard disc diffusion antimicrobial susceptibility tests

The influence of disc content on the zone sizes produced was investigated using standard disc diffusion antimicrobial agent susceptibility test protocols. Replicate measurements were made of the zones of inhibition produced by discs containing 2-64 μg of oxytetracycline and oxolinic acid against Escherichia coli, Aeromonas salmonicida and Vibrio parahaemolyticus. Five replicate data sets were obtained for each of the six organism-agent combinations. Analysis of these 30 data sets demonstrated that they all showed a very high correlation (mean r² = 0.98 ± 0.009) with the predictions of best-fit equations specifying a linear relationship between log₁₀ of the disc content (μg) and the resultant zone sizes (mm). The mean value of the slopes of the best-fit straight-line equations was 9.8 ± 1.8. The overall similarity of these values for the slopes suggested that the quantitative relationship between disc content and zone size was, at least to a first approximation, independent of the organism or the agent involved in the test.     

Breakpoints for disc diffusion susceptibility testing of bacteria associated with fish diseases: A review of current practice

A survey of the methods being employed to determine antimicrobial susceptibility of bacteria associated with aquaculture was performed on behalf of the Permanent Advisory Network for Diseases in Aquaculture. Thirty-two laboratories in 18 countries responded and 25 reported the breakpoints they used for disc diffusion assays applied to Group 1, non-fastidious organisms isolated from finfish. A total of 117 breakpoints were reported for assays in which the disc contents were those specified by the current standard protocols. Data on the source of these breakpoints and the confidence the laboratories had in them are presented. Overall there was a considerable variation in the breakpoints employed by different laboratories and this variation is discussed in terms of the inter-laboratory precision that can be expected from the application of disc diffusion protocols. This paper discusses the possible clinical significance of the variations in the breakpoints and, where there are available data, the extent to which those in use are consistent with breakpoints suggested by other approaches.The data presented in this paper represent a starting point for the movement towards harmonising breakpoints used in association with the standard disc diffusion protocols that have been proposed for susceptibility testing of bacteria associated with fish diseases.     

Effect of incubation temperature and time on the precision of data generated by antibiotic disc diffusion assays

The influence on the precision of disc diffusion data of the conditions under which the tests were performed was examined by analysing multilaboratory data sets generated after incubation at 35 °C for 18 h, at 28 °C for 24 h and 22 °C for 24 h and 48 h. Analyses of these data sets demonstrated that precision was significantly and progressively decreased as the test temperature was reduced from 35 to 22 °C. Analysis of the data obtained at 22 °C also showed the precision was inversely related to the time of incubation. Temperature and time related decreases in precision were not related to differences in the mean zone sizes of the data sets obtained under these test conditions. Analysis of the zone data obtained at 28 and 22 °C as single laboratory sets demonstrated that reductions of incubation temperature resulted in significant increases in both intralaboratory and interlaboratory variation. Increases in incubation time at 22 °C were, however, associated with statistically significant increases in interlaboratory variation but not with any significant increase in intralaboratory variation. The significance of these observations for the establishment of the acceptable limits of precision of data sets that can be used for the setting of valid epidemiological cut‐off values is discussed.