Background
Methods
General procedure
Plant material and extraction
Isolation and purification of bioactive compounds from the roots extract of A. adianthifolia
Antibacterial assays
Chemicals for antibacterial assays
Bacterial strains and culture media
Antibacterial testing
Results
Phytochemicals
Antibacterial activity
Bacterial strains | Tested samples, MIC and MBC in parenthesis (in μg/mL) | |||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Fractions | Fractions and sub-fractions | CHL | ||||||||||||
AARa | AARb | AARa1 | AARa2 | AARa3 | AARa4 | AARb1 | AARb2 | AARb11 | AARb12 | AARb13 | AAR14 | |||
E. coli
| AG100Atet | 1024(−) | 64(512) | 1024(−) | 512(−) | 512(−) | 1024(−) | 64(512) | 64(−) | 512(−) | 1024(−) | 64(−) | 256(−) | 32(256) |
AG102 | -(−) | 256(1024) | -(−) | 512(−) | 512(−) | 1024(−) | 128(−) | 512(−) | 1024(−) | -(−) | 128(1024) | 256(−) | 32(256) | |
ATCC8739 | 512(−) | 256(−) | -(−) | 512(−) | 512(1024) | 1024(−) | 16(256) | 64(512) | 64(1024) | 128(−) | 16(1024) | 16(1024) | 2(64) | |
ATCC 10536 | 1024(−) | 128(1024) | -(−) | 512(1024) | 512(1024) | 1024(−) | 32(1024) | 64(1024) | 128(−) | 256(−) | 32(−) | 64(−) | 2(32) | |
E. aerogenes
| ATCC13048 | 512(1024) | 128(1024) | -(−) | 256(1024) | 256(−) | – | 16(512) | 32(1024) | 128(−) | 512(1024) | 64(256) | 128(512) | 16(128) |
CM64 | 1024(−) | 256(1024) | -(−) | 256(512) | 512(1024) | – | 32(−) | 64(−) | 256(1024) | 512(−) | 32(128) | 256(1024) | 256(−) | |
EA27 | 512(−) | 8(128) | 1024(−) | 32(512) | 16(512) | 128(512) | 32(−) | 64(−) | 128(−) | -(−) | 32(1024) | 128(−) | 32(256) | |
EA289 | -(−) | 256(1024) | -(−) | -(−) | -(−) | -(−) | 32(−) | 64(−) | 512(−) | -(−) | 64(−) | 128(−) | 32(256) | |
K. pneumoniae
| ATCC11296 | 1024(−) | 256(−) | -(−) | 1024(−) | -(−) | -(−) | 16(−) | 64(−) | 128(−) | 512(−) | 32(512) | 32(256) | 32(256) |
KP55 | 256(−) | 128(−) | -(−) | 256(−) | 256(−) | -(−) | 16(1024) | 32(−) | 128(−) | 256(−) | 16(512) | 32(1024) | 64(256) | |
KP63 | 256(−) | 128(−) | 512(1024) | 256(−) | 512(−) | 1024(−) | 8(512) | 16(−) | 128(−) | 128(1024) | 8(128) | 128(−) | 32(256) | |
P.
stuartii
| ATCC29916 | -(−) | 128(1024) | 1024(−) | 512(−) | 512(1024) | -(−) | 128(−) | 256(−) | -(−) | -(−) | 256(−) | 512(−) | 64(256) |
NEA 16 | 1024(−) | 256(1024) | 1024(−) | -(−) | -(−) | -(−) | 128(−) | 256(−) | -(−) | -(−) | 128(1024) | 256(−) | 64(256) | |
P. aeruginosa
| PA01 | 1024(−) | 256(−) | -(−) | 512(−) | 512(−) | -(−) | 64(−) | 64(−) | 256(−) | 512(−) | 32(−) | 64(−) | 64(−) |
PA124 | -(−) | 256(−) | -(−) | 512(−) | 512(−) | -(−) | -(−) | -(−) | -(−) | -(−) | 1024(−) | -(−) | 256(−) |
Bacterial strains | Compounds, MIC and MBC in parenthesis (in μg/mL) | |||||
---|---|---|---|---|---|---|
2+ 3 | 4 | 5 | 6 | 7 | CHL | |
E. coli
| ||||||
AG102 | – | 128(−) | – | – | – | 32(−) |
ATCC8739 | – | 16(32) | – | – | – | 2(64) |
ATCC 10536 | 16(32) | 16(32) | – | 32(−) | 64(−) | 2(32) |
E. aerogenes
| ||||||
ATCC13048 | 128 (−) | 128(−) | 128(−) | 128(−) | 128(−) | 16(−) |
EA27 | – | – | – | – | – | 32(−) |
K. pneumoniae
| ||||||
ATCC11296 | – | – | 128(−) | – | – | 32(−) |
KP55 | 32(64) | – | 128(−) | 128 (−) | 128(−) | 64(−) |
P. stuartii |
–
| |||||
ATCC29916 | 64(128) | – | 64(128) | 128 (−) | – | 64(−) |
NEA 16 | – | – | – | – | – | 64(−) |
P. aeruginosa
| ||||||
PA01 | 16(64) | 2(64) | – | – | – | 64(−) |
PA124 | – | 128(−) | – | – | – | – |
Influence of the bacterial efflux pumps on the activity of the tested phytochemicals
Bacterial strains | Tested samples, MIC alone, MIC in the present of PAβN (μg/mL), and ameliorating factor (FA) | ||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
2 + 3 | 4 | 5 | 6 | 7 | CHL | ||||||||||||||
MIC | +PAβN | FA | MIC | +PAβN | FA | MIC | +PAβN | FA | MIC | +PAβN | FA | MIC | +PAβN | FA | MIC | +PAβN | FA | ||
E. coli
| AG102 | – | 128 | > 1 | 128 | 64 | 2 | – | – | – | – | – | – | – | – | – | 32 | 4 | 8 |
ATCC10536 | 16 | 4 | 4 | 16 | 8 | 2 | – | 128 | > 1 | 32 | 8 | 4 | 64 | 8 | 8 | 2 | < 1 | < 2 | |
E. aerogenes
| ATCC13048 | 128 | 64 | 2 | 128 | 32 | 4 | 128 | 64 | 2 | 128 | 64 | 2 | 128 | 32 | 4 | 16 | 8 | 2 |
EA27 | – | 128 | > 1 | – | 128 | > 1 | – | – | – | – | – | – | – | – | – | 32 | 16 | 2 | |
K. pneumoniae
| ATCC11296 | – | 128 | > 1 | – | 128 | > 1 | 128 | 32 | 4 | – | – | – | – | – | – | 32 | 8 | 4 |
KP55 | 32 | 8 | 4 | – | 128 | > 1 | 128 | 64 | 2 | 128 | 128 | 1 | 128 | 16 | 8 | 64 | 32 | 2 | |
P. stuartii
| ATCC29916 | 64 | 2 | 32 | – | 16 | > 8 | 64 | 16 | 4 | 128 | 32 | 4 | – | – | – | 64 | 8 | 8 |
NEA16 | – | 8 | > 16 | – | 16 | > 8 | – | 128 | > 1 | – | 128 | > 1 | – | 128 | > 1 | 64 | 16 | 4 | |
P. aeruginosa
| PA01 | 16 | 8 | 2 | 2 | < 1 | > 2 | – | – | – | – | – | – | – | – | – | 64 | 8 | 8 |
PA124 | – | 2 | > 64 | 128 | 64 | 2 | – | 128 | > 1 | – | – | – | – | – | – | 256 | 16 | 16 |
Potentiating effect of phytochemicals
Antibioticsa | Bacterial strainsb, MIC (μg/mL) of antibiotics in the absence and presence of compound 2 + 3 | ||||||||
---|---|---|---|---|---|---|---|---|---|
Compounds concentration | PA124 | KP55 | ATCC11296 | EA27 | ATCC13048 | AG102 | ATCC10536 | PBSS (%) | |
CIP | 0 | 2 | 0.5 | 0.5 | 0.5 | 4 | 2 | 0.125 | |
CMI/2 | 0.5(0.25)S | 0.5(1)I | 0.25 (0.5)S | 0.25 (0.5)S | 0.125(0.031)S | 0.5(0.25)S | 0.125 (1)I | (5/7) 71.42% | |
CMI/4 | 0.5(0.25)S | 0.5 (1)I | 0.25 (0.5)S | 0.5 (1)I | 0.5 (0.125)S | 1 (0.5)S | 0.125 (1)I | (4/7) 57.14% | |
ERY | 0 | > 32 | 4 | > 32 | > 32 | > 32 | 16 | 16 | |
CMI/2 | 32(0.5)S | 2(0.5)S | 4(< 0.125)S | 32 (0.5)S | 32 (0.5)S | 8 (0.5)S | 8 (0.5)S | (7/7) 100% | |
CMI/4 | 32(0.5)S | 2(0.5)S | 4(< 0.125)S | 32 (0.5)S | 32 (0.5)S | 8 (0.5)S | 8 (0.5)S | (7/7) 100% | |
GEN | 0 | > 4 | 2 | > 4 | 4 | 4 | > 4 | 4 | |
CMI/2 | 4(0.5)S | 1(0.5)S | 0.125(< 0.031)S | 4(1)I | 2(0.5)S | 4 (0.5)S | 2(0.5)S | (6/7) 85.71% | |
CMI/4 | 4(0.5)S | 1(0.5)S | 0.125(< 0.031)S | 4(1)I | 2(0.5)S | 4 (0.5)S | 2(0.5)S | (6/7) 85.71% | |
KAN | 0 | 0.5 | 2 | 4 | 4 | 16 | 16 | 4 | |
CMI/2 | < 0.125(0.25)S | 2(1)I | 2(0.5)S | 4(1)I | 16(1)I | 8 (0.5)S | 4(1)I | (3/7) 42.85% | |
CMI/4 | < 0.125(0.25)S | 2(1)I | 2(0.5)S | 4(1)I | 16(1)I | 8 (0.5)S | 4(1)I | (3/7) 42.85% | |
NOR | 0 | > 16 | 16 | 1 | 16 | 16 | 2 | 1 | |
CMI/2 | < 0.125(0.007)S | 8(0.5)S | 1(1)I | 4(0.25)S | 2(0.125)S | 2 (1)I | 1 (1)I | (4/7) 57.14% | |
CMI/4 | < 0.125(0.007)S | 8(0.5)S | 1(1)I | 4(0.25)S | 4(0.25)S | 2 (1)I | 1 (1)I | (4/7) 57.14% | |
STR | 0 | > 32 | > 32 | > 32 | > 32 | > 32 | > 32 | > 32 | |
CMI/2 | 32(0.5)S | 16(< 0.5)S | 32(0.5)S | 16(< 0.5)S | 32(0.5)S | 16(< 0.5)S | 2(< 0.062)S | (7/7) 100% | |
CMI/4 | 32(0.5)S | 32(0.5)S | 32(0.5)S | 16(< 0.5)S | 32(0.5)S | 16(< 0.5)S | 4(< 0.125)S | (7/7) 100% | |
TET | 0 | 8 | 0.125 | > 16 | > 16 | > 16 | > 16 | 16 | |
CMI/2 | 4(0.5)S | 0.0625(0.5)S | 8(< 0.5)S | 16(0.5)S | 4(< 0.25)S | 1(< 0.062)S | 0.125(0.007)S | (7/7) 100% | |
CMI/4 | 4(0.5)S | 0.0625(0.5)S | 8(< 0.5)S | 16(0.5)S | 4(< 0.25)S | 1(< 0.062)S | 0.125(0.007)S | (7/7) 100% |
Antibioticsa | Bacterial strainsb, MIC (μg/mL) of antibiotics in the absence and presence of compound 4 | ||||||||
---|---|---|---|---|---|---|---|---|---|
Compounds concentration | PA124 | KP55 | ATCC11296 | EA27 | ATCC13048 | AG102 | ATCC10536 | PBSS (%) | |
CIP | 0 | 2 | 0.5 | 0.5 | 0.5 | 4 | 2 | 0.125 | |
CMI/2 | 2 (1)I | 0.5(1)I | 0.25(0.5)S | 0.25(0.5)S | 0.5(0.125)S | 0.5(0.25)S | 0.125(1)I | (4/7) 57.14% | |
CMI/4 | 2 (1)I | 0.5(1)I | 0.25(0.5)S | 0.25(0.5)S | 0.5(0.125)S | 0.5(0.25)S | 0.125(1)I | (4/7) 57.14% | |
ERY | 0 | > 32 | 4 | > 32 | > 32 | > 32 | 16 | 16 | |
CMI/2 | 32 (0.5)S | 2(0.5)S | 4(< 0.125)S | 32 (0.5)S | 4(< 0.125)S | 4 (0.25)S | 8 (0.5)S | (7/7) 100% | |
CMI/4 | 32 (0.5)S | 2(0.5)S | 4(< 0.125)S | 32 (0.5)S | 8(< 0.25)S | 4 (0.25)S | 8 (0.5)S | (7/7) 100% | |
GEN | 0 | > 4 | 2 | > 4 | 4 | 4 | > 4 | 4 | |
CMI/2 | 4(0.5)S | 1(0.5)S | 0.0625(< 0.015)S | 1 (0.25)S | 2(0.5)S | 4(0.5)S | 0.5(0.125)S | (7/7) 100% | |
CMI/4 | 4(0.5)S | 2(1)I | 0.0625(< 0.015)S | 1 (0.25)S | (0.5)S | 4(0.5)S | 2(0.5)S | (6/7) 85.71% | |
KAN | 0 | 0.5 | 2 | 4 | 4 | 16 | 16 | 4 | |
CMI/2 | < 0.125(0.25)S | 1(0.5)S | 2(0.5)S | 4(1)I | 16(1)I | 16(1)I | 4(1)I | (3/7) 42.85% | |
CMI/4 | < 0.125(0.25)S | 2(1)I | 2(0.5)S | 4(1)I | 16(1)I | 16(1)I | 4(1)I | (2/7) 28.57% | |
NOR | 0 | > 16 | 16 | 1 | 16 | 16 | 2 | 1 | |
CMI/2 | 8(< 0.5)S | 2(0.125)S | 0.5(0.5)S | 4(0.25)S | 8(0.5)S | 2(1)I | 0.5(0.5)S | (6/7) 85.71% | |
CMI/4 | 8(< 0.5)S | 2(0.125)S | 0.5(0.5)S | 8(0.5)S | 8(0.5)S | 2(1)I | 0.5(0.5)S | (6/7) 85.71% | |
STR | 0 | > 32 | > 32 | > 32 | > 32 | > 32 | > 32 | > 32 | |
CMI/2 | 32(0.5)S | 32(0.5)S | 32(0.5)S | 16(< 0.5)S | 32(0.5)S | 16(< 0.5)S | 2(< 0.062)S | (7/7) 100% | |
CMI/4 | 32(0.5)S | 32(0.5)S | 32(0.5)S | 16(< 0.5)S | 32(0.5)S | 16(< 0.5)S | 2(< 0.062)S | (7/7) 100% | |
TET | 0 | 8 | 0.125 | > 16 | > 16 | > 16 | > 16 | 16 | |
CMI/2 | 4(0.5)S | 0.0625(0.5)S | 0.5(< 0.031)S | 4(< 0.25)S | 0.5(< 0.031)S | 2(< 0.125)S | 0.125(0.007)S | (7/7) 100% | |
CMI/4 | 4(0.5)S | 0.0625(0.5)S | 0.5(< 0.031)S | 8(< 0.5)S | 0.5(< 0.031)S | 2(< 0.125)S | 0.125(0.007)S | (7/7) 100% |