Table 1.

Susceptibilities to β-lactam antibiotics of S. pneumoniae transformants obtained with S. mitis B6 DNA

Strain or transformantaSelective antibioticb (μg/ml)PBP changedcMIC (μg/ml)
CefotaximeOxacillinBenzylpenicillinPiperacillin
S. pneumoniaeR60.020.080.020.04
S. mitisB6501205064
R6T-CC (0.2)2x0.2–0.250.2–0.50.03<0.2
R6T3-C C (0.2)2x0.2–0.250.2–0.50.03<0.2
R6T-CCC (2)2x, 2a1.5–2   0.50.02<0.2
R6T2-CC C (2)2x, 2a1.5–2   0.50.02<0.2
R6T-CCCC (6)2x, 2a, 1a 12–16  1–20.05–0.06<0.2
R6T5-CCC C (6)2x, 2a, 1a, 1b 12–16  0.5–1  0.05<0.2
R6T-CCCOO (200)2x, 2a, 1a, 1b, 2b 25–30   60–80 1–32–4
R6T-CCCO2b O (20)2x, 2a, 1a, 1b, 2b 25–35  601–22–4
R6T-CCCBB (10)2x, 2a, 1a, 1b, 2b50 80–1004016
R6T-CCPP (0.1)2x, 2a, 2b1–2  4–160.05–0.120.5–4   
R6T-PP (0.1)2b0.020.10.020.1–0.15
  • a Transformants were obtained with chromosomal DNA or in the case of R6T-CCCO2bwith the cloned PBP2b gene of S. mitis B6. Between three and eight transformants were tested. Boldface indicates the transformants used as recipients in successive transformations. Thus, R6T5-CCC is a third-step transformant obtained after three successive selections with increasing concentrations of cefotaxime and is the parental strain of (i) R6T-CCCO transformants obtained via oxacillin selection with chromosomal S. mitisB6 DNA, (ii) R6T-CCCO2b (oxacillin selection by using cloned PBP2b of S. mitis B6 as donor), and (iii) R6T-CCCB (transformation with chromosomal DNA and selection with benzylpenicillin).

  • b Antibiotics used for selection are listed in the order of the transformation steps used. The concentrations used at the last selection steps are indicated. C, cefotaxime; P, piperacillin; O, oxacillin; B, benzylpenicillin.

  • c 2x, PBP2x; 2a, PBP2a; 1a, PBP1a; 1b, PBP1b; 2b, PBP2b.