TABLE 1.

Quantification of germination and sporulationa

StrainIntestine sectionTime pointRNA (μg/g)DNA (μg/g)Minimum no. of cellsb% of inoculumcMaximum no. of cellsb% of inoculumc
PY79Jejunum (rrnO-tetC)12
1810.6 ± 3.479.4 ± 6.2(2.4 ± 0.7) × 108 1.20(1 ± 0.3) × 109 5
241.0 ± 0.411.4 ± 1.1(2.3 ± 0.9) × 107 0.12(9.6 ± 3.8) × 107 0.48
300.09 ± 0.040.6 ± 0.09(2.1 ± 0.9) × 106 0.01(8.6 ± 3.8) × 106 0.043
Ileum (cotB-tetC)302.0 ± 0.710.4 ± 1.9(4.6 ± 1.5) × 107 NA(1.9 ± 0.7) × 108 NA
360.6 ± 0.12.7 ± 0.4(1.4 ± 0.2) × 107 NA(5.8 ± 1) × 107 NA
480.03 ± 0.0080.15 ± 0.02(6.8 ± 1.8) × 105 NA(2.9 ± 0.8) × 106 NA
Large intestine (cotB-tetC)300.6 ± 0.12.6 ± 0.3(1.4 ± 0.2) × 107 NA(5.8 ± 1) × 107 NA
360.009 ± 0.0060.05 ± 0.008(2.1 ± 1.3) × 105 NA(8.8 ± 5.8) × 105 NA
480.007 ± 0.0010.03 ± 0.007(1.6 ± 0.2) × 105 NA(6.8 ± 1) × 105 NA
HU58Jejunum (rrnO-tetC)127.6 ± 1.562.7 ± 6.0(1.7 ± 0.3) × 108 0.85(7.3 ± 1.4) × 108 3.65
188.6 ± 2.870.8 ± 6.2(2.0 ± 0.6) × 108 1.00(8.3 ± 2.7) × 108 4.15
240.7 ± 0.35.9 ± 1(1.6 ± 0.7) × 107 0.08(6.8 ± 3) × 107 0.34
300.05 ± 0.020.5 ± 0.08(1.1 ± 0.5) × 106 0.006(4.8 ± 1.9) × 106 0.024
Ileum (cotB-tetC)301.6 ± 0.68.2 ± 1(3.6 ± 1.4) × 107 NA(1.5 ± 0.6) × 108 NA
360.9 ± 0.54.2 ± 0.7(2.1 ± 1.1) × 107 NA(8.8 ± 4.8) × 107 NA
480.07 ± 0.010.4 ± 0.07(1.6 ± 0.2) × 106 NA(6.8 ± 1) × 106 NA
Large intestine (cotB-tetC)300.6 ± 0.13.2 ± 0.3(1.4 ± 0.2) × 107 NA(5.8 ± 1) × 107 NA
360.03 ± 0.0080.18 ± 0.015(6.8 ± 1.8) × 105 NA(2.9 ± 0.8) × 106 NA
480.01 ± 0.0060.06 ± 0.008(2.3 ± 1.4) × 105 NA(9.6 ± 5.8) × 105 NA
  • a Samples of intestinal tissue from mice dosed with suspensions of spores expressing rrnO-tetC and cotB-tetC were removed, and total DNA and RNA were recovered as described in Materials and Methods and illustrated in Figure 3. RNA and DNA in jejunum samples were quantified for germination-specific gene expression using rrnO-tetC primers and ileum and large intestine samples for sporulation-specific gene expression with cotB-tetC primers. Using either RT-PCR for B. subtilis-specific RNA or PCR for B. subtilis-specific DNA, the quantity of each nucleic acid was determined by competitor and regression analysis and expressed in μg/g of sampled tissue. NA, not applicable. For PY79, the strain DL291 was used, and for HU58, strains BT17 and BT47 were used. A dash (—) indicates that no detectable PCR signal was apparent.

  • b Based on in vitro analysis of DNA extracted from cells growing in LB medium (see the supplemental material), the number of cells was approximated. Two estimates are given, a minimum estimate of the number cells based on 109 cells containing 10 chromosomes/cell and 44 μg DNA and a maximum estimate from 109 cells carrying 2 chromosomes/cell and containing 10 μg DNA.

  • c For samples showing germination, the number of cells based on DNA quantifications is expressed as a percentage of the original spore inoculum. Minimum and maximum estimated values are given.