- The ability of Streptococcus diacetilactis to inhibit a variety of
food-borne pathogens and spoilage organisms in milk and broth cultures
was demonstrated. Test organisms inhibited included Pseudomonas
and Alcaligenes species, Eschericia coli, Salmonella, Staphylococcus
aureus, Clostridium perfringens and Vibrio parahaemolyticus.
In general, approximately 99.0% and 99.9% inhibition was
observed in milk and broth cultures, respectively. Streptococcus
lactis, Streptococcus cremoris, Pediococcus cerevisiae and Lactobacillus
plantarum also inhibited S. aureus in lactic broth.
Possible practical applications of the observed inhibition were
examined. S. diacetilactis prevented proteolysis in milk at 7. 5C by
Pseudomonas fluorescens. S. aureus was inhibited greater than 99%
in vanilla cream filling, ham sandwich spread, chicken gravy, soy
milk and ground beef stored at 25C for 24 hr. Development of the
Gram negative flora of ground beef was also inhibited greater than
99% after storage at 7.5C for 7 days.
The mechanism of inhibition of S. aureus in lactic broth was
examined with emphasis on the role of pH changes and acid production
by S. diacetilactis. S. aureus did not grow in cell-free culture supernatants
of S. diacetilactis grown in modified lactic broth (final pH
4.3 - 4.7). However, good growth was evident when the pH was
adjusted towards neutrality. When the amounts of formic, acetic and
lactic acids produced by S. diacetilactis were quantitatively determined
and added to lactic broth, a similar pH dependent inhibition
was observed. This inhibition was not as marked as that observed
with cultures or cell-free supernatants of S. diacetilactis, suggesting
that factors other than acid production were involved. Nutrient depletion
and hydrogen peroxide did not contribute to the inhibition in this
Enterotoxin B synthesis by S. aureus S6 and S. aureus ATCC
14458 was studied in Brain Heart Infusion broth (BHI) and N-Z Amine
NAK broth (NAK). Toxin yields by S. aureus S6 were approximately
180 μg /ml and 100 μg/m1 in NAK and BHI, respectively at 30C and
150 rpm. Yields for strain 14458 were significantly lower. When
the initial inoculum of S. aureus S6 was 4 x 10⁵ cells/ml, the addition
of 0.5% glucose or 0.5% maltose initially repressed both toxin synthesis and the pH rise associated with toxin synthesis; however,
after 72 hr of incubation, the pH rise and toxin produced were the
same as occurred without added carbohydrate. Furthermore, addition
of maltose, but not glucose, reduced the toxin yield of S. aureus
ATCC 14458 by about 80% at 72 hr.
When S. diacetilactis (initial inoculum 1 x 10⁷ cells/ml) was
added to BHI containing 0. 5% glucose or 0. 5% maltose, no toxin synthesis
or pH rise was observed when the initial inoculum of S. aureus
was 4 x 10⁵ cells/ml. In plain BHI, no toxin was observed under
these associative growth conditions and the pH remained constant at
6.6. Inhibition of toxin production in this system could be partially
reversed by the addition of 0. 5% sodium pyruvate, sodium acetate or
When the initial inoculum of S. aureus was 4 x 10⁸ cells/ml,
only partial or no inhibition of toxin production in the presence of S.
diacetilactis (inoc ulum 1 x 10⁷ c ell s /m1) was observed. Substantial
inhibition by S. diacetilactis was observed at this concentration when
the medium was BHI + 0. 5% maltose but not when BHI + 0. 5% glucose
was used. Inhibition of toxin production but not growth was evident in
NAK + 1% glucose. Sodium citrate addition inhibited both growth and
toxin production by S. aureus S6.
The toxin yields of the S. aureus strains were greatly reduced
when static incubation was used. Inhibition by S. diacetilactis was
readily demonstrated under these conditions.
Three commercially available starter cultures, Lactacel,
Lactacel MC and Lactacel DS, were examined for their ability to control
the growth of S. aureus during a simulated beaker sausage fermentation
at 21C, 30C and 37C. Although differences between cultures
were evident, all three gave greater than 99% inhibition after
50 hr of incubation at 30C and 37C. Inhibition was somewhat reduced
(average of 97% at 50 hr) at 21C. Chemical acidulation with gluconodelta-
lactone (0.75%) plus citric acid (0. 1 %) yielded good initial control
in a similar process. However, on extended incubation up to
50 hr good growth of the pathogen occurred and the inhibition was
only slight or none at this time for samples incubated at 30C and 37C;
at 21C inhibition was approximately 97%. A combination of starter
cultures and chemical acidulation gave approximately 99.99% inhibition
of S. aureus at the three temperatures after 50 hr of incubation.
Possible applications of these findings to the food industry are
discussed. A greater role for the lactic acid bacteria in the area of
food-safety is suggested.