Negative and positive control ranges in the bacterial reverse mutation test: JEMS/BMS collaborative study

Abstract A large-scale study was conducted by multiple laboratories affiliated with the Japanese Environmental Mutagen Society and the Bacterial Mutagenicity Study Group to investigate possible proficiency indicators for the bacterial reverse mutation test with a preincubation procedure. Approximately 30 laboratories generated negative and positive control count data and dose-response curves of the positive control articles for the bacterial reverse mutation test, with assays conducted annually from 2013 to 2016. Overall, the majority of the negative and positive control counts for Salmonella Typhimurium strains TA100, TA1535, TA98, and TA1537, and Escherichia coli strain WP2uvrA, with and without S9 mix, were within the range of the means ±2× standard deviation. The negative counts were normally distributed (strains TA100, TA98, and WP2uvrA) or followed Poisson distribution (strains TA1535 and TA1537), and the positive control counts for all strains were approximately normally distributed. In addition, the distribution of the negative and positive control counts was relatively constant over the 4 years. The number of revertant colonies increased in a dose-dependent linear or exponential fashion up to the recommended doses for the respective positive control articles in Japan. These data are valuable for determining the acceptance criteria and an estimation of the laboratory proficiency for the bacterial reverse mutation test

Additional file 4: of Negative and positive control ranges in the bacterial reverse mutation test: JEMS/BMS collaborative study

Figure S3. Dose-response curves of revertant Salmonella Typhimurium strain TA1535 colonies following treatment with SA in the absence of S9 mix (a), or with 2AA in the presence of S9 mix (b). Individual dose-response curves were generated using results produced by each participating laboratory in 2016 (different colors indicate different laboratories). The doses tested were 0.125, 0.25, and 0.5 μg/plate for SA, and 0.5, 1.0, and 2.0 μg/plate for 2AA. (ODP 411 kb

Additional file 8: of Negative and positive control ranges in the bacterial reverse mutation test: JEMS/BMS collaborative study

Figure S7. Comparison of the negative and positive control counts for each strain with or without S9 mix. The doses shown in Figs. 6, 7, 8, 9, and 10 were used for comparison. The R2 values indicate the correlation coefficients of the linear regression lines. (ODP 62 kb

Additional file 6: of Negative and positive control ranges in the bacterial reverse mutation test: JEMS/BMS collaborative study

Figure S5. Dose-response curves of revertant Escherichia coli strain WP2uvrA colonies following treatment with AF-2 in the absence of S9 mix (a), or with 2AA in the presence of S9 mix (b). Individual dose-response curves were generated using results produced by each participating laboratory in 2016 (different colors indicate different laboratories). The doses tested were 0.0025, 0.005, and 0.01 μg/plate for AF-2, and 2.5, 5.0, and 10 μg/plate for 2AA. (ODP 342 kb

Additional file 3: of Negative and positive control ranges in the bacterial reverse mutation test: JEMS/BMS collaborative study

Figure S2. Dose-response curves of revertant Salmonella Typhimurium strain TA98 colonies following treatment with AF-2 in the absence of S9 mix (a), or treatment with 2AA in the presence of S9 mix (b). Individual dose-response curves were generated using results produced by each participating laboratory in 2016 (different colors indicate different laboratories). The doses tested were 0.025, 0.05, and 0.1 μg/plate for AF-2, and 0.125, 0.25, and 0.5 μg/plate for 2AA. (ODP 434 kb

Additional file 5: of Negative and positive control ranges in the bacterial reverse mutation test: JEMS/BMS collaborative study

Figure S4. Dose-response curves of revertant Salmonella Typhimurium strain TA1537 colonies following treatment with 9AA in the absence of S9 mix (a), or with 2AA in the presence of S9 mix (b). Individual dose-response curves were generated using results produced by each participating laboratory in 2016 (different colors indicate different laboratories). The doses tested were 20, 40, and 80 μg/plate for 9AA, and 0.5, 1.0, and 2.0 μg/plate for 2AA. (ODP 343 kb

Additional file 2: of Negative and positive control ranges in the bacterial reverse mutation test: JEMS/BMS collaborative study

Figure S1. Dose-response curves of revertant Salmonella Typhimurium strain TA100 colonies following treatment with AF-2 in the absence of S9 mix (a), or with 2AA in the presence of S9 mix (b). Individual dose-response curves were generated using results produced by each participating laboratory in 2016 (different colors indicate different laboratories). The doses tested were 0.0025, 0.005, and 0.01 μg/plate for AF-2, and 0.25, 0.5, and 1.0 μg/plate for 2AA. (ODP 423 kb

Additional file 1: of Negative and positive control ranges in the bacterial reverse mutation test: JEMS/BMS collaborative study

Table S1. Positive control articles and their doses used in this study (DOCX 18 kb