Characterization of Heat- resistant Fungus, Neosartorya pseudofischeri Using Proteomic Approach

Spoilage of pasteurized and canned fruit products caused by heat- resistant molds has been reported. These fungi usually contaminate fruits on or near the ground and survive heat treatments used for fruit processing resulting in growing and spoiling the products during storage at room temperature. The objectives of this study were to investigate the response of heat- resistant fungus, Neosartorya pseudofischeri isolated from the contaminated blueberry juice to various heat treatments, determine D-values and elucidate the mechanism(s) of fungal heat-resistance. 30 days old heat- resistant fungal spores were collected and inoculated into the blueberry juice and then subjected to heat treatments at different temperature for different duration. Mold counts were conducted and survivor curves were plotted. D-values were determined and compared using ANOVA. A quantitative proteomic approach employing stable isotope labeling by peptide demethylation was used to investigate the intracellular protein changes of N. pseudofischeri ascospores subjected to heat treatment at 93°C for 0, 1 or 8 min. The D-values of N. pseudofischeri at 85, 90, 93, 96 and 99 °C were 53.8, 12.2, 2.03, 0.98 and 0.42 min, respectively. In total, 811 proteins were identified and quantified and among them, 417 non-redundant proteins were classified into 16 functional categories while 150 significantly changed proteins were grouped into four clusters based on their quantitative changes. Decreased proteins were mainly involved in the central carbon metabolism, heat stress responses, reactive oxygen intermediates elimination and translation events. These proteins were stored during the spore formation under short stress condition, and their decrease was in agreement with the reduction of ascospores found following the heat treatment. A group of proteins in relation to toxicant degradation and antibiotic neutralization, which link to environmental adaptability and fungal heat-resistance were also identified. The results may be used by blueberry processors to prevent losses due to spoilage caused by the heat- resistant molds and establish appropriate thermal process schedules for fruit products. This study is the first report for proteomics on N. pseudofischeri that lays the foundation for further investigations on other heat- resistant molds using targeted quantitative studies.