TABLE 3.

Antibiotic susceptibilities and growth rates of M. smegmatis mutant strains with single and double 23S rRNA mutations^a

M. smegmatis straind	LZD MIC (μg ml−1)	LZD-nucleotide proximitye	Doubling time (h)	MIC (μg ml−1) of:
				CHL	CLI	VAL
SZ558 (parent)	2	NM	3.3 ± 0.1	32	16	32
Wild type	2	NM	3.5 ± 0.1	32	16-32	32
A2058G	2	C	3.5 ± 0.3	32	>1,028	64
G2032C	4	3	3.6 ± 0.6b	32	128	128
A2453U	4	2	3.9 ± 0.1	32	32	64
C2499U	2	3	4.1 ± 0.2	32	16	32
A2503G*	8	1	4.6 ± 0.1	128	8	32
G2505A*	16	1	6.4 ± 0.3	128	16	32
C2571G	4	X	4.0 ± 0.1	32	16-32	32
G2576U	64	2	9.1 ± 0.2	256	64	32
C2612A	4	X	4.2 ± 0.3	64	16	32
C2499A↓	2	3	3.8 ± 0.1	32	16	64
G2032A-C2499A	8	3-3	4.4 ± 0.1	64	16	128
G2032A↕	2	3	3.7 ± 0.3	32	32	128
G2032A-U2504G	64	3-1	>10c	256	16	128
U2504G*↕	8	1	5.8 ± 0.1	512	16-32	64
C2055A-U2504G	32	2-1	>10c	128	16-32	64
C2055A↕	0.5	2	3.8 ± 0.2	32	32	64
C2055A-A2572U	64	2-2	10.2 ± 0.6	64	512	128
A2572U↑	4	2	3.6 ± 0.3	8	16	128

^aMICs that differ from the unmutated strain sensitivity by 4-fold or more are indicated in bold type.

^bThe large standard deviation in the measured growth rates is due to cell clumping.

^cExact growth rate measurements could not be obtained due to cell clumping.

^dAsterisks indicate nucleotide positions that are interacting directly with linezolid. Arrows point to the corresponding double mutants.

^eThe positions of the mutated nucleotides relative to that of the bound linezolid drug are indicated, where 1, 2, and 3 represent the first, second, and third nucleotide layers around linezolid and X represents nucleotides outside of these three layers (see Fig. 1); C, control mutation; NM, no mutation. LZD, linezolid; CHL, chloramphenicol; CLI, clindamycin; VAL, valnemulin.