This study investigates the effect of citral on growth and on the occurrence of sublethal damage in Listeria innocua Serovar 6a (CECT 910) and Listeria monocytogenes Serovar 4b (CECT 4032) cells that were exposed to citral as a natural antimicrobial agent. Two initial inoculum concentrations were considered in this investigation: 102 and 106 cfu/mL. Citral exhibited antilisterial activity against L. innocua and L. monocytogenes, and the observed effects were dependent on the concentration of citral present in the culture medium (0, 0.150 and 0.250 μL/mL) (p ≤ 0.05). L. innocua had a shorter lag phase than L. monocytogenes, and the two species had nearly identical maximum specific growth rates. These results indicate that L. innocua could be used as surrogate for L. monocytogenes when testing the effects of this antimicrobial. Significant differences in the lag phase and growth rate were observed between the small and large inoculum concentration (p ≤ 0.05). Citral-treated L. innocua and L. monocytogenes that were recovered on selective medium (i.e., TSA-YE-SC) had a shorter lag phase and a higher maximum specific growth rate than cells that were recovered on non-selective medium (i.e., TSA-YE) (p ≤ 0.05). This result suggests that damage occurs at sublethal concentrations of citral.

No abstract

An in-depth study was conducted in order to extend the storage life of a liquid whole egg-skim milk (LWE-SM) mixed beverage to enhance its safety and the safety of related beverages. Bacillus cereus vegetative cells (1 x 10(8) colony-forming units [CFU]/mL) were inoculated in LWE-SM beverages with or without natural antimicrobial supplements: flavonol rich-cocoa powder (cocoanOX 12%, CCX) (700 ppm), vanillin (700 ppm), anise (700 ppm), and cinnamon (700 ppm). B. cereus cells were maintained at 10 degrees C for 10 days in the different beverages to test the bacteriostatic or inhibitory effect of the aforementioned ingredients. Beverages were treated with high hydrostatic pressure (HHP) technology and stored at 10 degrees C for 15 days after treatment. All natural antimicrobials reduced the micro(max) values and increased the lag phase time of B. cereus, and Gompertz growth curves showed different inhibitory effects depending on the substance. The maximum inhibitory effect (1.330 log cycle reduction) was achieved in LWE-SM-cinnamon-supplemented beverage. The maximum inactivation achieved by HHP in LWE-SM beverage was a reduction of around 3.89 +/- 0.25 log cycles at 300 MPa for 12 minutes. When supplemented beverages were treated under the same conditions, enhanced inactivation levels were achieved. This increased inactivation can be attributed to a synergistic effect when the LWE-SM was supplemented with flavonol-rich cocoa powder, cinnamon, and vanillin. The maximum synergistic effect was observed in LWE-SM-CCX-supplemented beverage. During the refrigerated storage of B. cereus HHP-treated cells in beverages to which antimicrobials had been added, the inhibitory effect was dependent on the previously applied pressure level.

Significance and Impact of the Study: Phytochemicals citral and carvacrol potentiate antibiotic activity of erythromycin, bacitracin and colistin by reducing the MIC values of cultured Listeria monocytogenes and Listeria innocua. This effect in reducing the MIC values of the antibiotics tested in both microorganisms was increased when natural antimicrobials were combined. This finding indicated that the combination among terpenes and antibiotic may contribute in reducing the required dosage of antibiotics due to the possible effect of terpenes on permeation barrier of the micro-organism cell membrane. AbstractThe aim of this study was to evaluate the antibiotic susceptibility of Listeria innocua (L. innocua) and Listeria monocytogenes (L. monocytogenes) cells in the presence of citral and carvacrol at sublethal concentrations in an agar medium. The presence of terpenes in the L. monocytogenes and L. innocua culture medium provided a reduction in the minimal inhibitory concentration (MIC) of all the antibiotics tested. These effects were dependent on the concentration of terpenes present in the culture medium. The combination of citral and carvacrol potentiated antibiotic activity by reducing the MIC values of bacitracin and colistin from 32Á0 and 128Á0 lg ml À1 to 1Á0 and 2Á0 lg ml À1 , respectively. Thus, both Listeria species became more susceptible to these drugs. In this way, the colistin and bacitracin resistance of L. monocytogenes and L. innocua was reversed in the presence of terpenes. Results obtained in this study show that the phytochemicals citral and carvacrol potentiate antibiotic activity, reducing the MIC values of cultured L. monocytogenes and L. innocua.

The main goal of this work was to study the bacterial adaptive responses to antibiotics induced by sublethal concentration of citral on first-and second-generation cells of Listeria monocytogenes serovar 4b (CECT 4032) and Salmonella enterica serovar Typhimurium (CECT 443). The first-generation cells were not pretreated with citral, while the second-generation cells were obtained from cells previously exposed to citral during 5 h. The trials were conducted at 37°C. The presence of citral in the culture medium and the antibiotic strips resulted in a reduced minimum inhibitory concentration (MIC) for the first-generation cells of Listeria monocytogenes serovar 4b and Salmonella Typhimurium. This result was observed for almost all the antibiotics, compared with the same microorganisms of the control group (without citral), which could represent an additive effect. For Listeria serovar 4b, the second-generation cells of the test group maintained the same susceptibility to antibiotics compared with cells in the control group and in the test group of the first generation. The second-generation cells of the control group indicated that the Salmonella Typhimurium maintained the same sensitivity to the antibiotics tested compared with the first generation of this group, except in the case of erythromycin, which exhibited an increased MIC value. With respect to the second-generation cells of Salmonella Typhimurium, the presence of citral determined a decrease in the antibiotic susceptibility for almost all of the antibiotics, except colistin, compared with the first-generation of the test group, which can be seen by increase of MIC values. In conclusion, the presence of citral in the culture medium of Listeria 4b and Salmonella Typhimurium increased the antibiotic susceptibility of the first generations, while we observed an increase in antibiotic resistance in the second generation of Salmonella Typhimurium.