Proline Increases Pigment Production to Improve Oxidative Stress Tolerance and Biocontrol Ability of Metschnikowia citriensis

Abstract

Utilizing antagonistic yeasts is a promising approach for managing postharvest decay of fruits. However, it is well established that various severe stresses encountered in the environment and production process cause the intracellular reactive oxygen species (ROS) accumulation in yeast cells, resulting in cell damage and loss of vitality. Here, proline has been shown to function as a cell protectant and inducer of biofilm formation able to increase the oxidative stress tolerance and the biocontrol ability of the antagonistic yeast Metschnikowia citriensis . Addition of proline to M. citriensis cells induced a significant rise in superoxide dismutase (SOD) and catalase (CAT) activity in the early and late stages of oxidative stress, respectively, and increased the maroon pigment production that directly reduced intracellular iron content and indirectly diminished intracellular ROS levels and thus inhibited ROS- and iron-induced apoptosis. Treating cells with iron chelator tropolone yielded similar results. Pigment production induced by proline also enhanced the capability of biofilm formation of M. citriensis . These results suggested an important role for pigment of M. citriensis in response to oxidative stress. The abilities of proline to scavenge intracellular ROS and inhibit apoptosis, increase pigment production, and promote biofilm formation contribute to the improvements in oxidative stress tolerance and biocontrol efficacy of M. citriensis .

Metschnikowia pulcherrima and Related Pulcherrimin-Producing Yeasts: Fuzzy Species Boundaries and Complex Antimicrobial Antagonism

“…These experiments were carried out with Metschnikowia isolates whose exact taxonomic affiliation could not be determined because their D1/D2 sequences differed from those of all type strains of the M. pulcherrima clade [ 6 ]. Correlation and causative relationship between iron depletion (pulcherrimin production) and antagonism was later observed in many other pulcherrimin-producing Metschnikowia strains, including the type strains of the species (e.g., [ 7 , 8 , 10 , 11 , 21 , 71 , 72 , 73 ]).…”

“…Microorganisms can exert harmful effects by secretion of enzymes that damage the walls or other components of the cells of other microorganisms. Several strains of the M. pulcherrima clade were found able to secrete extracellular lytic enzymes, such as chitinase and glucanase (e.g., [ 10 , 11 , 75 , 76 , 77 , 100 , 101 ]) but the significance of this secretion is disputable in certain cases. For example, the antagonistic strain MACH1 secreted chitinase in various media including apple juice [ 76 ].…”

“…In contrast, Parafati et al [ 10 ] detected neither β-1,3-glucanase nor chitinase secretion in a different collection of Metschnikowia strains. Even closely related strains can differ in the production of extracellularly measurable chitinase activity: FL01 showed chitinase activity but no extracellular enzyme activity was detected in the culture of FL02 [ 11 ]. In the cells of the strain 277 the chitinase gene CHI was found upregulated not only by the presence of P. digitatum hyphae but also by the presence of grapefruit peel tissue [ 61 ].…”

“…In a different study, the increased pulcherrimin production of the strain FL01was accompanied with increased biofilm formation [ 11 ]. This observation led to the hypothesis that the secreted pulcherriminic acid protects the plant tissues against the pathogens not only by sequestering the iron but also by promoting the yeast cells to form films which hold up the fungal invasion [ 11 , 73 ]. For the applicability of an antagonistic strain to bioprotection, it is also important that its cells adhere to the surface of the plant to be protected.…”

“…Due to the large fatty globules in their chlamydospores (“pulcherrima cells”), the strains are outstanding candidates for low-cost lipid production (reviewed in [ 4 ]). Their most intensively studied property is the strong antimicrobial activity ( Figure 1 A–C) (e.g., [ 5 , 6 , 7 , 8 , 9 , 10 , 11 , 12 ]). Since the mechanisms underlying their antagonistic effect are not associated with the production of toxic compounds, these strains can safely be used as bioprotective agents to curb the invasion of pathogenic and rotting (saprophytic) microorganisms ( Figure 1 D) and/or additives in food technologies to modulate the dynamics of microbial populations.…”