Antifungal activity is a relatively common characteristic among bacteria, conferring an ecological advantage in environments which support the growth of a mixed bacterial and fungal flora. This activity has been detected by using a variety of in vitro methods and although the chemical basis for this activity has been elucidated in many cases, in some it has not, even though antifungal activity has been demonstrated. This activity has significance in four areas: development of therapeutic antifungal drugs, development of plant protection agents and suppression of fungal colonization: proliferation within the human body resulting in modification of the pattern of certain human clinical infections and reduction in the efficiency of isolation of fungal pathogens from clinical specimens. Fluorescent pseudomonads representing the group of PGPR can promote growth and suppress plant pathogens by multiple mechanisms. Their applicability as bio-controlling agents has drawn wide attention because of production of secondary metabolites such as siderophores, antibiotics, volatile compounds, HCN, enzymes and phytohormones. Pseudomonas fluorescens is considered as a biological bio-controlling agent against various plant related diseases including root diseases. The use of bacterial remedies in the fight with pests and various mycoses encountered at agricultural plants represents a major interest. Some of the most popular remedies with entomo- and phytopathogenic activity are obtained from bacteria of the genus Bacillus and Pseudomonas. In this regard, the strains of microorganisms that are used in biotechnology are representing a commercial value and the problem of maintaining as longer as possible of their biosynthetic proprieties and of those that are economic valuable are permanently in the attention of scientists. The conservation of microorganisms and of their properties requires the use of efficient methods of preservation and a continuous monitoring of the effectiveness of these methods. A special interest aroused the maintenance of the antifungal activity of bacterial strains of Pseudomonas aurantiaca CNMN-PsB-08, Pseudomonas aureofaciens CNMN-PsB-07 and Bacillus cereus var. fluorescens CNMN-BB-07 that are stored in the NCNM (National Collection of Nonpathogenic Microorganism) of the Institute of Microbiology and Biotechnology of the ASM, after a long period of time as a result of the lyophilization. As pathogenic cultures, the following fungal strains were used: Fusarium oxysporum, Fusarium solani, Alternaria alternata, Botrytis cinerea and Aspergillus niger. The results of investigations undertaken on determining the antifungal activity of the strains of Ps. aurantiaca CNMN-PsB-08, Ps. aureofaciens CNMN-PsB-07 and B. cereus var. fluorescens CNMN-BB-07 over the strains of fungal pathogens after their freeze-drying storage in the protective environment of Na succinate + 12% sucrose during the 3 and 6 years, proved that together with the extension of the storage period increases also the antifungal activity. Thus, the collected data are demonstrating that after 6 years of conservation of strains of Ps. aurantiaca CNMN-PsB-08 and B. cereus var. fluorescens CNMN-BB-07 the antifungal activity on micromycetes F. oxysporum and F. solani is increasing so, that the diameter of the inhibition zone is increasing by 5.0 – 4.7 mm and 6.3 – 7.7 mm, respectively, from the preservation period of 3 years. The antifungal activity of all isolates has increased together with the increase of the storage period of 3 to 6 years towards the micromycetes A. alternata and B. cinerea which also are active pathogens of the crop plants. In this case, we can observe that the diameter of the inhibition zone of the strain Ps. aurantiaca CNMN-PsB-08 towards the A. alternata and B. cinerea is growing by 8.6 and 5.3 mm, respectively. Unlike this, the strains of Ps. aureofaciens CNMN-PsB-07 and B. cereus var. fluorescens CNMN-BB-07 are possessing antifungal activity also over the A. niger culture, thus the diameter of the inhibition zone is increasing from 12.3 to 19.0 mm and from 14.3 to 15.0 mm, respectively. The most significant increase of the antifungal activity was determined at the strain Ps. aureofaciens CNMN-PsB-07 over the micromicetes B. cinerea, where the inhibition zone was increasing together with the storage period from 3 to 6 years with 2.03 times. The use of these bacterial species in biological control would allow increasing the plant resistance to pathogens. Thus, the achievement of some efficient conservation methods that would allow the maintenance or the increase of antifungal properties can be finalized with the development of some technological equipment for the attainment of the active biological remedies, efficient in the agricultural usage as an alternative to chemical fungicides.