TABLE 6.
Cumulative mutations decreasing susceptibility to LYS228 or ATM in E. coli over 20 serial passages
| Strain | Gene mutated (aa substitution) | MIC (μg/ml) |
|||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|
| LYS228 (+ AVI) | ATM (+ AVI) | CAZ | CAZ/AVI | IPM | MEM | LVX | CIP | TGC | CHL | ||
| NB27001 | Parent | 0.125 (0.125) | 0.125 (0.125) | 0.25 | 0.25 | 0.06 | 0.03 | 0.03 | 0.008 | 0.25 | 8 |
| NB27001-JBC0001a | Selected by LYS228; ftsI(Q536L), baeR(L22Q), envZ(L84Q), acrD(R629H), sucB(P27T), rfaI(Y112C) | 32 (8) | 16 (16) | 8 | 8 | 0.06 | 0.03 | 0.03 | 0.015 | 0.125 | 8 |
| NB27001-JWM0095 | ftsI(Q536L) | 0.5 (0.25) | 0.5 (0.25) | 2 | 1 | 0.125 | 0.03 | 0.015 | 0.008 | 0.125 | 4 |
| NB27001-JWM0094 | baeR(L22Q) | 1 (0.5) | 1 (1) | 1 | 1 | 0.06 | 0.03 | 0.03 | 0.015 | 0.125 | 4 |
| NB27001-JWM0096 | envZ(L84Q) | 0.5 (0.5) | 0.5 (1) | 1 | 1 | 0.125 | 0.06 | 0.03 | 0.015 | 0.25 | 4 |
| NB27001-JWM0093 | acrD(R629H) | 0.25 (0.25) | 0.25 (0.5) | 0.25 | 0.5 | 0.06 | 0.03 | 0.03 | 0.015 | 0.125 | 8 |
| NB27001-JRW0009 | sucB(P27T) | 0.125 (0.25) | 0.5 (0.25) | 0.5 | 1 | 0.06 | 0.03 | 0.03 | 0.015 | 0.5 | 8 |
| NB27001-JRW0008 | rfaI(Y112C) | 0.5 (0.5) | 1 (0.5) | 0.5 | 1 | 0.06 | 0.03 | 0.03 | 0.008 | 0.5 | 8 |
| NB27001-JRW0027 | baeR(L22Q), acrD(R629H) | 1 (1) | 2 (1) | 2 | 2 | 0.06 | 0.06 | 0.015 | 0.015 | 0.25 | 4 |
| NB27001-JBC0006a | Selected by ATM; ftsI(A413V), baeR(P111Q), rseA(W33R), rfaH(T72P), yhbM(W24*), frameshifts in acrB, irhA, and yghB; | 16 (4) | 16 (8) | 4 | 2 | 0.06 | 0.06 | ≤0.008 | 0.008 | 0.125 | 2 |
| NB27001-JWM0101 | ftsI(A413V) | 1 (0.25) | 1 (0.5) | 0.25 | 0.5 | 0.06 | 0.03 | 0.015 | 0.008 | 0.25 | 4 |
| NB27001-JWK0180 | ftsI (YRIK insertion) | 0.5 (0.5) | 1 (1) | 1 | 2 | 0.06 | 0.03 | 0.015 | 0.008 | 0.125 | 4 |
| NB27001-JWK0181 | ftsI (YRIN insertion) | 0.5 (1) | 0.5 (1) | 1 | 2 | 0.06 | 0.03 | 0.015 | 0.008 | 0.06 | 8 |
a
Representative isolate of five that were genome sequenced and had the identical set of mutations. All mutations were confirmed by PCR and sequencing. Only ftsI(A413V) from NB27001-JBC006 was tested for impact on susceptibility to LYS228. Increased susceptibility to CHL in NB27001-JBC0006 is consistent with the identified frameshift mutation in acrB. Each individual mutation identified from the LYS228 serial passage was engineered into the genome of strain NB27001.