Table 3. The in vivo effect of mutations in the conserved region of MdsA^a in the presence of wild-type MdsB and TolC without MdsC.

Background	ATCC14028SΔacrABΔmdsABC::Cm5
MdsA protein	MIC (µg/mL)b
	Crystal violet	Methylene blue	Acriflavine	Rhodamine 6G
Nonec	4	32	16	4
MdsA-WT	16	256	32	64
MdsA-R135D	8	64	16	32
MdsA-L139D	4	32	16	8
MdsA-S146D	16	256	32	64

^aThe in vivo effect of each mutation in the conserved region of MdsA was determined by measuring the resistance of the ATCC14028SΔacrABΔmdsABC::Cm5 strain to several substrates. MdsA and its variants were expressed from pMdsAB2, pMdsAB2 MdsA-R135D, pMdsAB2 MdsA-L139D, and pMdsAB2 MdsA-S146D.

^bAll MIC measurements were done in triplicate. The concentrations of crystal violet and acriflavine used to determine the MICs were 0, 1, 2, 4, 8, 16, and 32 µg/mL. The concentrations of rhodamine 6G and methylene blue used to determine the MICs were 0, 4, 8, 16, 32, 64, 128, and 256 µg/mL.

^cNone, strain carrying the empty vector pKAN6B instead of an pMdsAB2 variant.