Isolation and identification of a marine killer yeast strain YF07b and cloning of the gene encoding killer toxin from the yeast

It was found that the marine yeast strain YF07b could secrete a large amount of killer toxin against a pathogenic yeast strain WCY which could cause milky disease in Portunus trituberculcttus. The marine yeast strain YF07b was identified to be Pichia anomala according to the results of routine yeast identification and 18S rDNA and ITS sequences. The gene encoding killer toxin in the marine yeast strain YF07b was amplified by PCR technology. After sequencing, the results show that an open reading frame, consisting of 1 281 bp, encoded a presumed protein of 427 amino acids. The sequence of the cloned gene was found to have 99% match with that of the gene encoding killer toxin in Pichia anomalas strain K. A signal peptide including 17 amino acids appeared in the N-terminal domain of the killer toxin. Therefore, the mature protein consisted of 410 amino acids, its molecular mass was estimated to be 47.4 ku and its isoelctronic point was 4.5.

Antifungal Activity of Lactic Acid Bacteria, Isolated from Bulgarian Wheat and Rye Flour

The economic losses and the health hazards of the mycotoxins produced by spoilage fungi are the main concerns of the food industry. The spoilage of bakery products by fungi is more common in countries with a high humidity and temperature. About 5-10% of food production is spoiled by the growth of yeast and fungi in food materials. Similarly, in Western Europe, the growth of the spoilage fungi of bread is estimated to reach more than 200 million Euros per year. The history conditions of the food can be a major factor in determining any fungal spoilage--for example, stored and processed foods are more sensitive to spoilage when compared with fresh and prepared foods. Lactic acid bacteria isolated from Bulgarian wheat and rye flour were used in the present study to check their antifungal properties against pathogenic yeast and fungi imperfecta using standard disc diffusion method in vitro. A broad spectrum of antifungal activity of the six newly identified as L. plantarum strains e Tsl, Ts2, Ts3,Ts4 and Ts5, and L. helveticus Ts6 was estimated. Our in vitro studies were performed with wheat and rye sourdough, in order to simulate a real product and to assess the bio-protective potential of the tested lactobacilli. The used test-cultures are representatives of carcinogenic, toxigenic, deteriorative and allergenic fungi from the genera .4spergillus and Penicillium. The all tested strains completely suppress the growth of against C. glabrata 72. Strains L. plantarum Tsl and Ts3 completely suppress the growth against S. cerevisae. While, in the sample with L. plantarum strains e Tsl, Ts2, Ts3,Ts4 and Ts5, and L. helveticus Ts6, a retarded and weak growth of A. niger and P. claviforme was observed. However, the spore germination and the colony growth started only on the fifth day of the mould lactobacilli co-cultivation, which also should be considered as a good result. In this study six isolates Tsl,Ts2, Ts3, Ts4, Ts5 and Ts6, from the traditional Bulgarian wheat and rye flour have been identified as L. plantarum and L. helveticus and characterized as cultures with promising antifungal activity. Obtained results from the combined molecular identification （16S rRNA gene sequencing） approach contribute to give new data on the microbial biodiversity of this not well-studied niche. The antifungal activity of our new isolates, identified as L. plantarum and L. helveticus, seems to be a promising advantage of these six strains, suggesting their potential applications in different food technologies. However, more experiments have to be conducted to clarify the nature and the mechanisms of the reported antifungal activity and they are still in progress. The combination of dairy origin and strong inhibitory activity of the lactobacillus strains is a prerequisite for their possible application as starters and/or bioprotective antifungal adjuncts.