This study screened 24 strains of Oenococcus oeni against foodborne pathogens like E. coli O157:H7, Salmonella enteritidis, and Listeria monocytogenes. Most strains inhibited the growth of one or more pathogens. Four strains were selected for further characterization of their antimicrobial compounds. Tests determined the antimicrobial activity was due to organic acids rather than proteases, catalase, or other potential compounds. Future work will identify the specific antimicrobial compounds and evaluate O. oeni's effectiveness in controlling pathogens in various food applications.

1. B-1
Part 2 (Chapter 3)
Growth Inhibition of Foodborne Pathogens
by Oenococcus oeni

2. B-2
Introduction
LAB are good for foodborne pathogen control:
acids, antimicrobial compounds, GRAS, no
negative impacts

3. B-3
Aims of this study
1. Screening O. oeni against foodborne pathogen
2. Characterize antimicrobial compounds
Foodborne
pathogensO. oeni

4. B-4
Cultures
vs
24 strains
O. oeni
E. coli O157:H7
L. monocytogenes
Salm. enteritidis

5. B-5
250
C
8 days
Inoculating
producers
Indicator
s
370
C
1 to 3 days
Screening

6. B-6
Medium-MLB for O. oeni
pH was adjusted to 4.5 by adding KOH
MLB with 1.5% agar
TSB with 0.75% agar
Tryptic Soy Broth (TSB) for pathogens
Pancreatic digest of casein, Enzymatic digest of soybean
meal ,sodium chloride 5 g l-1

7. B-7
CH3, Figure 3.1
Most strains of O. oeni
were capable of
inhibiting E. coli O157:H7
Results

8. B-8
CH3, Figure 3.2
Most strains of O. oeni
were capable of inhibiting
Salm. enteritidis

9. B-9
Inhibition spectrum of O. oeni against foodborne pathogens
Indicator
Producer
E. coli O157:H7 43895
Salm. enteritidis L. monocytogenes
104035
THKOe01 / / +
THKOe02 / + +
THKOe04 + + +
THKOe05 / / +
THKOe06 + + +
THKOe07 + + +
THKOe08 + + +
THKOe09 + + +
THKOe10 + + +
THKOe11 + + +
THKOe12 + + +
THKOe13 + + +
THKOe14 + + +
THKOe15 + + +
THKOe16 - / +
THKOe17 / / +
THKOe18 + + +
THKOe19 + + +
THKOe20 / + +
THKOe21 + + +
THKOe22 + + -
THKOe23 + + +
THKOe24 + + +
THKOe25 + + +
+, inhibition; -, no inhibition; /, inhibition zone was hazy
CH3, Table 3.1

10. B-10
E. coli O157:H7--> inhibited by 18 strains
Salm. enteritidis --> inhibited by 20 strains
L. monocytogenes --> inhibited by 23 strains
THKOe04, 06, 10 and 23 were selected for
characterizing antimicrobial compounds.

11. B-11
For identifying antimicrobial compounds:
Protease
Catalase
Buffer solution
Analysis of
potential compounds
Preparation of
compounds
Spot-on-lawn
method
Well
diffusion
method

12. B-12
CH3, Table 3.2 Sensitivity tests
Producer and treatment
Indicator
E. coli O157:H7 Salm. enteritidis L. monocytogenes
THKOe04 Protease Type IX - - -
Protease Type XIII - - -
Protease Type XIV - - -
Protease Type XVIII - - -
Pronase E - - -
Savinase - - -
Catalase - - -
Buffer / - -
THKOe06 Protease Type IX - - -
Protease Type XIII - - -
Protease Type XIV - - -
Protease Type XVIII - - -
Pronase E - - -
Savinase - - -
Catalase - - -
Buffer + - /
THKOe10 Protease Type IX - - -
Protease Type XIII - - -
Protease Type XIV - - -
Protease Type XVIII - - -
Pronase E - - -
Savinase - - -
Catalase - - -
Buffer + + +
THKOe23 Protease Type IX - - -
Protease Type XIII - - -
Protease Type XIV - - -
Protease Type XVIII - - -
Pronase E - - -
Savinase - - -
Catalase - - -
Buffer / - -

13. B-13
Catalase --> negative
6 proteases --> negative
Buffer solution -->
1. THKOe06 vs E. coli O157H7 --> totally inactivated
2. THKOe10 vs three pathogens --> totally inactivated
==> due to acids
3. THKOe04 vs E. coli O157H7 --> partially inactivated
4. THKOe23 vs E. coli O157H7 --> partially inactivated
5. THKOe06 vs Listeria --> partially inactivated
==> multiple factors

14. B-14
For antimicrobial test with
known compounds

15. B-15
Single tests MLB, pH 4.1, 100 μl
Acetaldehyde: 10, 20, 50, 100, 200 and 500 mg l-1
(ND)
Diacetyl: 10, 20, 50, 100, 200 and 500 mg l-1
(ND)
Ethanol: 100, 250, 500 and 1000 mg l-1
(Max 466 mg l-1
)
Acetic acid: 1, 2, 5, and 10 g l-1
(Max 1.88 g l-1
)
Lactic acid: 5, 7, 14 and 21 g l-1
(7.00 g l-1
)
Combination tests
MLB with 500 mg l-1
ethanol, 2 g l-1
acetic acid and 7 g l-1
lactic
acid, pH 4.1.
With or without additions of 200 mg l-1
acetaldehyde and 20
mg l-1
diacetyl.
--> All negative on pathogens

16. B-16
Conclusion
1. This is the first report of using O. oeni for
foodborne pathogen control.
2. Antimicrobial activity can be due to acids and
other factors.

17. B-17
Future Work
1. Continue to identify antimicrobial compounds.
2. Application evaluation: pathogens control in
foods, salad, fresh or cooked meats, and
probiotics (like vitamin pills).
3. Evaluate the antimicrobial activity by O. oeni
along and with others antimicrobials (H2
O2
)
4. Try more pathogens e.g. Staphylococcus
aureus, Bacillus cereus.