Traditional sourdoughs in Bulgaria were almost extinct during the centralized food production system. However, a rapidly developing trend of sourdough revival in the country is setting the demand for increased production and use of commercial starter cultures. The selection of strains for such cultures is based on geographical specificity and beneficial technological properties. In this connection, the aim of this study was to isolate, identify and characterize lactic acid bacteria (LAB) and yeasts from typical Bulgarian sourdoughs for the selection of strains for commercial sourdough starter cultures. Twelve samples of typical Bulgarian sourdoughs were collected from different geographical locations. All samples were analyzed for pH, total titratable acidity and dry matter content. Enumeration of LAB and yeast was also carried out. Molecular identification by 16S rDNA sequence analysis was performed for 167 LAB isolates, and 106 yeast strains were identified by ITS1-5.8S-ITS2 rRNA gene partial sequence analysis. The LAB strains were characterized according to their amylolytic and proteolytic activity and acidification capacity, and 11 strains were selected for further testing of their antimicrobial properties. The strains with the most pronounced antibacterial and antifungal activity are listed as recommended candidates for the development of starter cultures for sourdoughs or other food products.

Grapes (Vitis vinifera L.) are an essential crop for fresh consumption and wine production. Vineyards are attacked by several economically important bacterial and fungal diseases that require regular pesticide treatment. Among them, Pseudomonas syringae pv. syringae (Ps. syringae) and Botrytis cinerea (B. cinerea) infections cause huge economic losses. The fresh fruit market has shifted to functional natural foodstuffs with clear health benefits and a reduced use of chemicals along the production chain. Lactic acid bacteria (LAB) have a biopreservative effect and are applied to ensure food safety in response to consumers’ demands. In the present study, the possibilities of using microorganisms with a potential antimicrobial effect against Ps. syringae and B. cinerea in the production of table grapes were investigated. LAB of the genus Lactiplantibacillus can be a natural antagonist of pathogenic bacteria and fungi by releasing lactic acid, acetic acid, ethanol, carbon dioxide and bacteriocins in the medium. The present study focuses on the characterization of nine Lactiplantibacillus plantarum (Lp. plantarum) strains isolated from spontaneously fermented sourdoughs. Species-specific PCR identified the isolated LAB for partial recA gene amplification with an amplicon size of 318 bp. RAPD-PCR analysis showed the intraspecific diversity of the individual strains. Thirteen plantaricin-like peptides (PlnA, PlnB, PlnC, PlnD, PlnEF, PlnG, PlnI, PlnJ, PlnK, PlnN, PlnNC8, PlnS, and PlnW) produced by isolated Lp. plantarum strains were detected by PCR with gene-specific primers. The key features for future industrial applications were their antimicrobial properties. The culture medium and cell-free supernatant (CFS) were used to establish in vitro antimicrobial activities of Lp. plantarum strains against Ps. syringae and B. cinerea, and inhibition of phytopathogen development was observed. The inhibitory effect of the CFS (cell-free supernatant) of all strains was assessed by infecting table grapes with these pathogens in in vivo experiments. Lp. plantarum Q4 showed the most effective suppression of the pathogens both in vitro and in vivo, which indicates its potential use as a biocontrol agent against berry rot and grey rot on grapes, caused by Ps. syringae and B. cinerea.

Exopolysaccharides (EPSs) produced by lactic acid bacteria present a particular interest for the food industry since they can be incorporated in foods via in situ production by selected starter cultures or applied as natural additives to improve the quality of various food products. In the present study, 43 strains were isolated from different plant-based fermented foods and identified by molecular methods. The species found were distinctively specific according to the food source. Only six Lactiplantibacillus plantarum strains, all isolated from sauerkraut, showed the ability to produce exopolysaccharide (EPS). The utilization of glucose, fructose and sucrose was explored with regard to EPS and biomass accumulation by the tested strains. Sucrose was clearly the best carbon source for EPS production by most of the strains, yielding up to 211.53 mg/L by strain Lactiplantibacillus plantarum ZE2, while biomass accumulation reached the highest levels in the glucose-based culture medium. Most strains produced similar levels of EPS with glucose and fructose, while fructose was utilized more poorly for biomass production, yielding about 50% of biomass compared to glucose for most strains. Composition analysis of the EPSs produced by strain Lactiplantibacillus plantarum ZE2 from glucose (EPS-1) and fructose (EPS-2) revealed that glucose (80–83 mol%) and protein (41% w/w) predominated in both analyzed EPSs. However, the yield of EPS-1 was twice higher than that of EPS-2, and differences in the levels of all detected sugars were found, which shows that even for the same strain, EPS yield and composition vary depending on the carbon source. These results may be the basis for the development of tailored EPS-producing starter cultures for food fermentations, as well as technologies for the production of EPS for various applications.

In this research, the biosynthetic and biocontrol potential of endophytic yeast to improve the growth and development of tobacco has been elucidated. Three yeast strains were enriched and isolated from different plant tissues. Partial sequence analysis of ITS5-5.8-ITS4 region of the nuclear ribosomal DNA with universal primers identified YD5, YE1, and YSW1 as Saccharomyces cerevisiae (S. cerevisiae), Zygosaccharomyces bailii (Z. bailii), and Saccharomyces kudriavzevii (S. kudriavzevii), respectively. When cultivated in a medium supplemented with 0.1% L-tryptophan, isolated yeast strains produced indole-3-acetic acid (IAA). The capacities of those strains to improve the mobility of phosphorus and synthesize siderophores has been proven. Their antimicrobial activities against several Solanaceae plant pathogenic fungi (Alternaria solani pathovar. tobacco, Rhizoctonia solani, and Fusarium solani pathovar. phaseoli) were determined. S. cerevisiae YD5, Z. bailii YE1, and S. kudriavzevii YSW1 inhibited the growth of all tested pathogens. Yeast strains were tested for endophytic colonization of tobacco by two different inoculation methods: soil drench (SD) and leaf spraying (LS). To establish colonization in the various tissues of tested tobacco (Nicotiana tabaccum L.) plants, samples were taken on the seventh, fourteenth, and twenty-first days after treatment (DAT), and explants were inoculated on yeast malt agar (YMA). Both techniques of inoculation showed a high frequency of colonization from 83.33% to 100%. To determine the effectiveness of the microbial endophytes, their effect on some physiological processes in the plant were analyzed, such as photosynthesis, stomatal conductivity, and transpiration intensity. The effect of single and double treatment with yeast inoculum on the development and biochemical parameters of tobacco was reported. Plants have the ability of structural and functional adaptation to stress effects of different natures. All treated plants had a higher content of photosynthetic pigments compared to the control. Photosynthesis is probably more intense, and growth stimulation has been observed. The chlorophyll a/b ratio remained similar, and the total chlorophyll/carotenoid ratio slightly increased as a result of elevated chlorophyll levels. The most significant stimulating effect was recorded in tobacco plants treated by foliar spraying with Z. bailii YE1 and S. cerevisiae YD5. In contrast, S. kudriavzevii YSW1 had a better effect when applied as a soil drench. Thus, S. cerevisiae YD5, Z. bailii YE1, and S. kudriavzevii YSW1 have a high potential to be used as a biocontrol agents in organic agriculture.

The present study investigated developing biological control agents against plant pathogens as an alternative to pesticides. The plant growth-promoting (PGP) and biocontrol potential of bacteria from the Bacillus genus is due to their ability to produce proteolytic and amylolytic enzymes, assist in the solubilization of phosphorus and zinc, and the production of siderophores. Cell culture and cell-free supernatant were used to investigate the antimicrobial activity of different Bacillus strains against the phytopathogenic fungus Fusarium graminearum in vitro. Fusarium graminearum is a fungus that causes plant disease, particularly in cereals like wheat and barley. As a result, significant suppression of the growth and development of this plant pathogen was observed. Plant growth-promoting activity manifested when the bacteria were applied alone and in combination. A single strain and combinations of two, three, and four strains of Bacillus were tested for their antimicrobial effects against Fusarium graminearum. The fluorescence spectroscopy results proved that the combination of Bacillus subtilis, Bacillus circulans, Bacillus megaterium, and Bacillus licheniformis showed the best stimulation of development, expressed as a comparative evaluation of the yield compared to the untreated control variant. The four strains showed their potential application as a biocontrol agent against Fusarium graminearum. The four Bacillus strains also can promote plant growth by affecting nutrition, root structure, and plant health, and they have the capacity to dissolve phosphates and zinc.

The accumulation of biowastes is one of the main concerns of modern society. One of the most environmentally friendly solutions to convert biowaste into a product is composting. Biowastes may contain unknown substances that are persistent in the final compost, thus contributing to soil contamination and salinization. The effectiveness of the composting process depends on the microbial communities involved, which is the number of investigations’ targets. The present work studied the bacterial diversity of mesophilic and thermophilic phases of composting developed in two different sites. The study was conducted through next-generation Illumina HiSeq sequencing and phylogenetic communities, revealing the dynamics and changes in specific mesophilic and thermophilic habitats of composting piles. The results showed a higher number of bacterial species in the mesophilic phase than in the thermophilic one, proved by the Shannon and Chao indices. In addition, the diversity of bacterial species expressed by the operational taxonomic units was much higher at the site of Harmanli than at the Yasno pole. Higher abundance was found of the genera Sphingobacterium, Sphingomonas, Paracoccus, Pseudomonas, and Halomonas in both studied sites. In the compost of Harmanli genera Streptomyces, Truepera, and Flavobacterium were found to be much more abundant compared to the compost of the Yasno pole. Finally, we conclude that the two plots show relatively significant differences in the diversity of bacteria during biowaste composting. Substantial differences were also observed between the mesophilic and thermophilic phases, with the first showing a significantly higher degree of species richness.