Impact of heat shock and osmotic/matric stress adaptation of yeasts biocontrol agents against Aspergillus Section Flavi

MONTEMARANI, A.; NESCI, A.; ETCHEVERRY, M.

Rotterdam

Conferencia; 7th conference of the World Mycotoxin Forum and XIIIth IUPAC International Symposium on Mycotoxins and Phycotoxins; 2012

Kluyveromyces spp. have biocontrol efficacy against Aspergillus Section Flavi in maize. Temperature and osmotic stress play an important role in biocontrol system. Ecological fitness, the ability of an organism to survive and replicate, is closely related to the performance of biocontrol agents; thus, enhancing stress tolerance may represent a strategy for improving efficacy. Heat shock treatment, characterized by the induction of a trehalose-6-phosphate synthase gene and other specific proteins, can enhance the ability of yeasts to withstand unfavorable environmental conditions, such as freeze drying, hydrostatic pressure, high temperature and oxidative stress. The aims of this study were (1) to determine the heat shock survival upon cells of yeast and (2) to determine the accumulation of endogenous compatible solutes of yeast cells. Kluyveromyces spp. L16 and L25 were initially grown in MSB [molasses (20 gL-1) + soy powder (10 gL-1)] and NYDB 75% [nutrient broth (6 gL-1) + dextrose (7,5 gL-1) + yeast extract (3,75 gL-1)] osmotically modified to 0.99, 0.97 and 0.95 aw using the non-ionic solute glucose. These media supported a rapid growth rate and a high biomass production. When cultures reached the stationary phase of growth, cells were harvested by centrifugation. The cells were incubated for 1 h in separate water baths set at 35, 40 and 45°C and later, a new count of viable cells was done on meal extract agar plates. Yeasts cells were grown in meal extract broth with aw modified with NaCl and glycerol and pH 3, 5 and 7, for the quantification of intracellular compatible solutes. Twenty five mg cell material was analyzed for intracellular solutes trehalose, glycerol, erythritol, arabitol, glucose and mannitol, and quantified by HPLC. The viability of L16 and L25 increased with increasing temperature. As unexpected, at 45°C cells showed an increasing count (12 log cfu ml-1). The manipulation of biocontrol agents L16 and L25, by changes in aw with different types of solutes, incubation times and pH affected the accumulation of compatible solutes. The solutes accumulated in higher proportion were glycerol and arabitol. Cells grown under unmodified aw condition showed accumulation of significant content of trehalose. The tolerance of both yeasts to water potential modifications and heat shock could increase the potential for biological control of Aspergillus Section Flavi in the storage of grains. Furthermore, these results could have important implications for optimizing and improving formulations.