Data were analyzed by one-way analysis of variance with a Tukey-Kramer multiple comparisons post-test. *, DNA per cell was significantly greater (P < 0.05) in exponentially growing B. burgdorferi B31 cultured

at 34°C in the presence of 6% rabbit serum than in stationary phase B. burgdorferi under any culture condition examined. **, RNA per cell was significantly lower (P < 0.05) in stationary phase B. burgdorferi B31 cultured at 23°C in the presence of 6% rabbit serum than in B. burgdorferi cultured at 34°C in the presence or absence of 6% rabbit serum. Figure 4 Detection of (p)ppGpp in B. burgdorferi B31 grown at 34°C in BSK-H in the presence (lane 1) or absence (lane 2) of 6% rabbit serum, or in BSK-H at 23°C in the presence of

6% rabbit serum (lane 3). Similar amounts of (p)ppGpp Sirolimus cell line were detected in cells under all three culture conditions. For calculation of DNA, RNA and protein on a per cell basis, data from washed exponential and stationary phase cells were analyzed separately (see legend to Figure HCS assay 3 for details). Since we could not obtain sufficient amounts of material for analysis of exponential growth at 23°C because of the relative insensitivity of colorimetric assays and high costs of large volumes of BSK-H culture medium, only the data from day 11 were used for this condition. At 34°C, there was significantly more DNA per cell in exponentially growing B. burgdorferi B31 cultures containing rabbit serum than at any of the other growth conditions (P < 0.05, one-way analysis of variance, Tukey-Kramer multiple comparison post-test) (Figure 3E). There was significantly less RNA per cell in stationary phase B. burgdorferi at 23°C than at 34°C (Figure 3F) (P < 0.05, one-way analysis of variance, Tukey-Kramer multiple comparison post-test). There was no significant

difference in protein per cell under any growth condition at any temperature (Figure 3G). Because precise correlation between corresponding points on growth curves for cultures growing at different rates is difficult, it was therefore still unclear whether rRNA levels were regulated by growth rate or growth phase in B. burgdorferi B31. Effect of growth rate and stringent response mafosfamide on accumulation of 16S and 23S rRNA in B. burgdorferi The apparent effect of growth rate on rRNA synthesis could be influenced by sampling B. burgdorferi that were in different growth phases at the two temperatures. Direct analysis of 16S and 23S rRNA levels in B. burgdorferi B31 grown in BSK-H containing serum at 34°C and 23°C revealed no difference in the levels of normalized 16S rRNA expression in cells grown at different temperatures when the cells were at similar points in the growth phase (Figure 5A).