Background
Methods
Bacterial strains and culture conditions
pH and bile tolerance tests
Antibiotic susceptibility testing
Preparation of pathogen strain inoculums
Antimicrobial activity of LAB “cell-free cultures supernatants”
Aggregative abilities of LAB strains
Cell cultures
Adhesive properties of LAB strains on Caco-2 cells
Invasion inhibition by interference studies with selected LAB strains and their combinations
Statistical analysis
Results
Strain-specific probiotic properties
Amp | Van | Gen | Kan | Strep | Ery | Clin | Tetra | Chlo | |
---|---|---|---|---|---|---|---|---|---|
B. bifidum W23 | 0.06 [2] | 1 [2] | 64 [64] | n.r. | 32 [128] | 0.25 [1] | 0.12 [1] | 2 [8] | 2 [4] |
L. acidophilus W37 | 0.094 [1] | 0.5 [2] | 1 [16] | 32 [64] | 3 [16] | 0.016 [1] | 0.094 [1] | 0.38 [4] | 1.5 [4] |
L. casei W56 | 0.5 [4] | n.r. | 4 [32] | 64 [64] | 32 [64] | 0.25 [1] | 0.12 [1] | 2 [4] | 8 [4] |
L. plantarum W21 | 0.5 [2] | n.r. | 8 [16] | 64 [64] | n.r. | 0.5 [1] | 2 [2] | 32 [32] | 4 [8] |
L. rhamnosus W71 | 4 [4] | n.r. | 2 [16] | 32 [64] | 8 [32] | 0.12 [1] | 0.5 [1] | 1 [8] | 4 [4] |
L. salivarius W24 | 1 [4] | n.r. | 2 [16] | 64 [64] | 32 [64] | 0.25 [1] | 0.25 [1] | 4 [8] | 4 [4] |
L. lactis W58 | 0.25 [2] | 0.25 [4] | 2 [32] | 8 [64] | 16 [32] | 0.12 [1] | 0.12 [1] | 1 [4] | 4 [8] |
Antimicrobial activity of CFCSs against food-borne pathogens
CFCSs | Inhibition zone (mm ± sd) | ||||
---|---|---|---|---|---|
S. enteritidis
ATCC 13076 |
L. monocytogenes
ATCC 7644 |
E. coli O157: H7 ATCC 35150 |
C. sakazakii
ATCC 29544 |
C. jejuni
ATCC 33291 | |
B. bifidum W23 | – | 10.1 ± 0.25 | 11.0 ± 0.35 | 12.1 ± 0.25 | 12.1 ± 0.32 |
L. salivarius W24 | 10.1 ± 0.32 | – | – | 10.2 ± 0.11 | 10.1 ± 0.28 |
L. acidophilus W37 | – | 11.1 ± 0.12 | – | – | 12.1 ± 0.36 |
L. casei W56 | – | 11.1 ± 0.51 | – | 10.2 ± 0.21 | 17.1 ± 0.21 |
L. lactis W58 | 11.0 ± 0.15 | 11.0 ± 0.27 | 12.0 ± 0.28 | 11.1 ± 0.15 | 10.5 ± 0.51 |
L. plantarum W21 | 10.1 ± 0.31 | 14.1 ± 0.34 | 10.0 ± 0.20 | 10.1 ± 0.28 | 13.5 ± 0.52 |
L. rhamnosus W71 | 10.1 ± 0.24 | 12.1 ± 0.31 | 11.0 ± 0.15 | 10.2 ± 0.25 | 17.2 ± 0.21 |
Positive control | 16.3 ± 0.53 (Gen) | 19.3 ± 0.63 (W) | 15.6 ± 0.64 (Gen) | 16.3 ± 0.52 (Strep) | 32 ± 0.51 (Gen) |
Aggregation abilities and adhesiveness of LAB strains
Auto-aggregation | Co-aggregation with pathogens | |||||
---|---|---|---|---|---|---|
S. enteritidis
ATCC 13076 |
L. monocytogenes
ATCC 7644 |
E. coli O157: H7 ATCC 35150 |
C. sakazakii
ATCC 51329 |
C. jejuni
ATCC 33291 | ||
B. bifidum W23 | 21.37 (±1.12) | 14.86 (±0.28) | 11.96 (±0.20) | 10.27 (±0.31) | 10.07 (±0.28) | 18.14 (±1.02) |
L. salivarius W24 | 19.33 (±1.81) | 6.82 (±0.20) | 8.33 (±0.12) | 10.31 (±0.21) | 7.06 (±0.22) | 15.84 (±0.32) |
L. acidophilus W37 | 15.90 (±1.20) | 14.2 (±0.19) | 8.35 (±0.11) | 3.85 (±0.01) | 9.52 (±0.19) | 4.33 (±0.02) |
L. casei W56 | 15.92 (±1.72) | 6.34 (±0.11) | 5.14 (±0.15) | 5.64 (±0.14) | 5.34 (±0.11) | 13.94 (±0.32) |
L. lactis W58 | 15.50 (±1.21) | 13.94 (±0.31) | 7.13 (±0.13) | 9.32 (±0.21) | 4.54 (±0.31) | 3.42 (±0.28) |
L. plantarum W21 | 15.03 (±1.61) | 16.79 (±0.35) | 14.62 (±0.31) | 12.01 (±0.24) | 4.72 (±0.34) | 13.00 (±0.21) |
L. rhamnosus W71 | 21.08 (±1.83) | 15.93 (±0.24) | 13.73 (±0.22) | 17.4 (±0.22) | 12.51 (±0.26) | 11.00 (±0.23) |
Interference of LAB strains on intestinal pathogens invasion ability
Strains and interference test | Single LAB strains | LAB combinations | |||||
---|---|---|---|---|---|---|---|
W23 | W24 | W71 | W23 + W24 | W23 + W71 | W24 + W71 | W23 + W24 + W71 | |
S. enteritidis ATCC 13076 | |||||||
Exclusion (%) | 27.92 | 31.82 | 57.14 | 55.19 | 80.52 | 74.68 | 68.83 |
Competition (%) | 20.13 | 45.45 | 47.40 | 62.99 | 82.47 | 53.25 | 80.52 |
L. monocytogenes ATCC 7644 | |||||||
Exclusion (%) | 76.94 | 90.78 | 86.16 | 97.23 | 95.39 | 97.23 | 95.85 |
Competition (%) | 90.78 | 90.78 | 95.39 | 97.23 | 99.08 | 98.16 | 98.16 |
E. coli O157:H7 ATCC 35150 | |||||||
Exclusion (%) | 88.11 | 77.95 | 66.14 | 93.70 | 93.70 | 92.13 | 93.70 |
Competition (%) | 65.35 | 49.61 | 52.76 | 94.49 | 78.74 | 85.83 | 77.95 |
C. sakazakii ATCC 29544 | |||||||
Exclusion (%) | 56.19 | 61.90 | 82.86 | 71.67 | 63.33 | 61.67 | 66.67 |
Competition (%) | 65.71 | 23.81 | 52.38 | 68.33 | 66.67 | 56.67 | 71.67 |
C. jejuni ATCC 33291 | |||||||
Exclusion (%) | 94.80 | 88.30 | 96.90 | 95.00 | 96.00 | 90.70 | 97.00 |
Competition (%) | 80.00 | 73.00 | 99.10 | 93.00 | 97.00 | 94.00 | 94.00 |