- November 22, 2013 at 1:58 am #112791
dvjorgeParticipantTopics: 283Replies: 1369
3.2. Fungicidal Activities of Lactobacilli
A microscopic approach was taken to visualize whether the probiotic lactobacilli are in fact able to kill the fungal cells. FUN-1 LIVE/DEAD staining following cocultures of the bacteria with C. albicans revealed that fungal cells lost metabolic activity in the presence of the lactobacilli and eventually were killed (Figure 4(a)). Semiquantitative analyses of C. albicans cell viability following exposure to low pH, lactic acid, and the bacterial strains used in this study revealed again that lactic acid at low pH and the probiotic strains GR-1 and RC-14 exert the most potent antifungal properties. The microplate-based assay system was used to determine cell viability by kinetic measurement of the intracellular conversion of the green fluorescent FUN-1 dye to red fluorescent intravacuolar structures. Only viable and metabolically active cells are able to carry out this conversion which can be monitored by determination of red/green fluorescence ratios in a fluorometer. Figure 4(b) shows the effect of MRS with lactic acid at pH 4.5 compared to MRS with HCl-adjusted pH 4.5 on C. albicans cell viability after overnight incubation. Presence of lactic acid under these conditions appears to affect fungal cell viability substantially more than just the low pH. The efficacy of the probiotic strains GR-1 and RC-14 to compete with C. albicans growth is also confirmed by the fluorometric viability assay results shown in Figure 4(c). Both strains generated the flattest red/green ratio curves indicating the strongest antagonistic effects on the fungi in this assay system. L. johnsonii PV016 appeared to have an intermediate effect on fungal cell viability while S. aureus only produced a minor reduction in the conversion rate from green to red fluorescence when compared to the C. albicans control (see Figure 4(c)). Overall, these viability assay results provide strong evidence that the probiotics are not only competing for nutrients and have fungistatic properties—under suitable conditions they can indeed exert fungicidal effects.
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