Evaluation of eliciting activity of peptidil prolyl cys/trans isomerase from Pseudonomas fluorescens encapsulated in sodium alginate regarding plant resistance to viral and fungal pahogens.

Use of chemical pesticides poses a threat for environment and human health, so green technologies of crop protection are of high demand. Some microbial proteins able to activate plant defense mechanisms and prevent the development of resistance in plant pathogens, may be good alternative to chemicals, but practical use of such elicitors is limited due to need to protect them against adverse environment prior their delivery to target receptors of plant cells. In this study we examined a possibility to encapsulate heat resistant FKBP-type peptidyl prolyl cis-trans isomerase (PPIase) from Pseudomonas fluorescens, which possesses a significant eliciting activity in relation to a range of plant pathogens, in sodium alginate microparticles and evaluated the stability of resulted complex under long-term UV irradiation and in the presence of proteinase K, as well as its eliciting activity in three different "plant-pathogen" models comparing to that of free PPIase. The obtained PPIase-containing microparticles consisted of 70% of sodium alginate, 20% of bovine serum albumin, and 10% of PPIase. In contrast to free PPIase, which lost its eliciting properties after 8-h UV treatment, encapsulated PPIase kept its eliciting ability unchanged; after being exposed to proteinase K, its eliciting ability twice exceeded that of free PPIase. Using "tobacco-TMV", "tobacco-Alternaria longipes", and "wheat-Stagonospora nodorum" model systems, we showed that encapsulation process did not influence on the eliciting activity of PPIase. In the case of the "wheat-S. nodorum" model system, we also observed a significant eliciting activity of alginate-albumin complex and almost doubled activity of encapsulated PPIase as compared to the free PPIase. As far as we know, this is the first observation of a synergistic interaction between alginate and other compound possessing any bioactive properties. The results of the study show some prospects for a PPIase use in agriculture.

New bacitracin-resistant nisin-producing strain of Lactococcus lactis and its physiological characterization.

Nisin A belonging to the class I bacteriocins and produced by Lactococcus lactis subsp. lactis is widely used in many countries as highly efficient and safe preservative preventing growth of undesirable bacteria in food products. Though this compound is efficient at very low concentrations, reduction of its manufacturing cost is still relevant problem. An increased nisin A production requires improved resistance of its producer to nisin. According to some studies, mechanisms of microbial resistance to nisin A and bacitracin have a similar basis, and the same transporters are used to export these antibiotics from cells. To obtain strains with improved growth rate and nisin A productivity, selection of spontaneous bacitracin-resistant L. lactis mutants followed by examination of their stability as well as physiological and fermentation characteristics was carried out. Spontaneous mutants were obtained by culturing of L. lactis VKPM B-2092 strain on selective bacitracin-containing agar medium. The obtained bacitracin-resistant strain FL-75 was characterized by accelerated growth rate, doubled biomass accumulation, and improved nisin A resistance. The nisin A productivity of FL-75 exceeded that of the parental strain by 25% reaching 8902 U/mL after 14-h cultivation. In addition, FL-75 was characterized by the improved resistance to oxidative stress that has never been reported earlier for bacitracin-resistant microorganisms. Based on the performed characterization of FL-75, we can consider it as a new independent strain promising for the industrial production of food and feed biopreservatives. Comparison of published data and the obtained results allowed us to suppose that the bacitracin resistance mutation in FL-75 is determined rather by an increased expression of a gene homologous to the bcrC gene of Bacillus sp. than by the activation of multidrug resistance mechanisms. The revealed resistance of FL-75 to bacitracin and oxidative stress can be regulated by a common transcription factor activating in response to various environmental stresses.

Effect of Various Compounds Blocking the Colony Pigmentation on the Aflatoxin B1 Production by Aspergillus flavus.

Aflatoxins and melanins are the products of a polyketide biosynthesis. In this study, the search of potential inhibitors of the aflatoxin B1 (AFB1) biosynthesis was performed among compounds blocking the pigmentation in fungi. Four compounds-three natural (thymol, 3-hydroxybenzaldehyde, compactin) and one synthetic (fluconazole)-were examined for their ability to block the pigmentation and AFB1 production in Aspergillus flavus. All compounds inhibited the mycelium pigmentation of a fungus growing on solid medium. At the same time, thymol, fluconazole, and 3-hydroxybenzaldehyde stimulated AFB1 accumulation in culture broth of A. flavus under submerged fermentation, whereas the addition of 2.5 µg/mL of compactin resulted in a 50× reduction in AFB1 production. Moreover, compactin also suppressed the sporulation of A. flavus on solid medium. In vivo treatment of corn and wheat grain with compactin (50 µg/g of grain) reduced the level of AFB1 accumulation 14 and 15 times, respectively. Further prospects of the compactin study as potential AFB1 inhibitor are discussed.

Strain Improvement of Streptomyces xanthochromogenes RIA 1098 for Enhanced Pravastatin Production at High Compactin Concentrations.

Pravastatin is one of the most popular cholesterol-lowering drugs. Its industrial production represents a two-stage process including the microbial production of compactin and its further biocatalytic conversion to pravastatin. To increase a conversion rate, a higher compactin content in fermentation medium should be used; however, high compactin concentrations inhibit microbial growth. Therefore, the improvement of the compactin resistance of a producer still remains a relevant problem. A multi-step random UV mutagenesis of a Streptomyces xanthochromogenes strain RIA 1098 and the further selection of high-yield compactin-resistant mutants have resulted in a highly productive compactin-resistant strain S 33-1. After the fermentation medium improvement, the maximum bioconversion rate of this strain has reached 91 % at the daily compactin dose equal to 1 g/L and still remained high (83 %) even at the doubled dose (2 g/L). A 1-year study of the mutant strain stability has proved a stable inheritance of its characteristics that provides this strain to be very promising for the pravastatin-producing industry.