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[ÇóÖú] International Journal of Food Microbiology

Refrigeration alone or in combination with other technologies represents the main tool used in the last decades
to preserve the freshness of black truffles. This is principally due to the delicateness and vulnerability of this
edible hypogeous fungus, so that other invasive preservation practices cannot be adopted. However, the pro?liferation of some microbial species during the cold storage still represents an unsolved problem. Pseudomonads
are among the main spoiler bacteria responsible for the deterioration of refrigerated black truffles. Their growth
ability at low temperatures requires the use of additional hurdles to prolong the shelf-life of truffles without
altering their major features. The use of natural compounds may represent an alternative system for the bio?control of this kind of product. Specifically, gallic acid (GA) is a phenolic acid naturally present in different
foods, whose effectiveness was in vitro demonstrated against Pseudomonas spp. In our study, we reported the
antimicrobial activity expressed by GA not only in vitro, using as target bacteria Pseudomonas putida DSMZ 291T
,
P. fluorescens DSMZ 50090T
, P. fragi DSMZ 3456T and Pseudomonas spp. P30-4, previously isolated from black
truffles, but also in situ on fresh black truffles stored at 4 ¡ãC for 28 days. Our results showed Minimum Inhibitory
Concentrations (MIC) of 2.5 mg/mL GA for all tested strains, except for P. fluorescens DSMZ 50090T
, having a
MIC corresponding to 5 mg/mL GA. The Minimum Bactericidal Concentration (MBC) was 10 mg/mL for all
strains. The analysis of kinetic parameters showed that the survival declined passing from 2.5 to 10 mg/mL GA
concentrations, with P. fluorescens confirmed to be the most resistant strain. Moreover, images obtained from
Scanning Electron Microscopy revealed that Pseudomonas cells were strongly injured by the treatment with GA at
2.5 mg/mL concentration, displaying visible pores on the cellular surfaces, absence of flagella and lysis with loss
of cytoplasmic material.
The storage test performed on fresh black truffles confirmed in situ the GA antimicrobial activity observed in
vitro, with a drastic reduction not only of Pseudomonas spp., but also of the other assessed microbial groups,
including Enterobacteriaceae and Eumycetes. Finally, sensory analysis established the absence of off-flavours and
the preservation of positive features in black truffles treated with 2.5 mg/mL GA and stored for 28 d at 4 ¡ãC. The
results obtained in this study suggest that GA is a potential biocontrol tool to decontaminate and preserve fresh
black truffles during refrigerated storage.

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