Saccharomyces paradoxus K66 Killer System Evidences Expanded Assortment of Helper and Satellite Viruses.

The Saccharomycetaceae yeast family recently became recognized for expanding of the repertoire of different dsRNA-based viruses, highlighting the need for understanding of their cross-dependence. We isolated the Saccharomyces paradoxus AML-15-66 killer strain from spontaneous fermentation of serviceberries and identified helper and satellite viruses of the family Totiviridae, which are responsible for the killing phenotype. The corresponding full dsRNA genomes of viruses have been cloned and sequenced. Sequence analysis of SpV-LA-66 identified it to be most similar to S. paradoxus LA-28 type viruses, while SpV-M66 was mostly similar to the SpV-M21 virus. Sequence and functional analysis revealed significant differences between the K66 and the K28 toxins. The structural organization of the K66 protein resembled those of the K1/K2 type toxins. The AML-15-66 strain possesses the most expressed killing property towards the K28 toxin-producing strain. A genetic screen performed on S. cerevisiae YKO library strains revealed 125 gene products important for the functioning of the S. paradoxus K66 toxin, with 85% of the discovered modulators shared with S. cerevisiae K2 or K1 toxins. Investigation of the K66 protein binding to cells and different polysaccharides implies the ß-1,6 glucans to be the primary receptors of S. paradoxus K66 toxin. For the first time, we demonstrated the coherent habitation of different types of helper and satellite viruses in a wild-type S. paradoxus strain.

Preparation and characterization of iron oxide magnetic nanoparticles functionalized by nisin.

Nisin is a known bacteriocin approved as a food additive for food preservation. It exhibits a wide spectrum antimicrobial activity against Gram-positive bacteria. Iron oxide magnetic nanoparticles were synthesized and characterized by X-ray diffraction method. A main part of iron oxide nanoparticles was found to be maghemite though a small quantity of magnetite could also be present. Magnetic nanoparticles were stabilized by citric, ascorbic, gallic or glucuronic acid coating. Stable iron oxide magnetic nanoparticles were functionalized by nisin using a simple and low cost adsorption method. Nisin loading was confirmed by FT-IR spectra, thermogravimetric analysis, dynamic light scattering and atomic force microscopy methods. Nisin-loaded iron oxide magnetic nanoparticles were stable at least six weeks as judged by the measurements of zeta-potential and hydrodynamic diameter. The antimicrobial activity of nisin-loaded iron oxide magnetic nanoparticles was demonstrated toward Gram-positive bacteria. Functionalized nanoparticles could therefore find the application as antimicrobials in innovative and emerging technologies based on the magnetic field.

Overcoming Antimicrobial Resistance in Bacteria Using Bioactive Magnetic Nanoparticles and Pulsed Electromagnetic Fields.

Nisin is a known bacteriocin, which exhibits a wide spectrum of antimicrobial activity, while commonly being inefficient against Gram-negative bacteria. In this work, we present a proof of concept of novel antimicrobial methodology using targeted magnetic nisin-loaded nano-carriers [iron oxide nanoparticles (NPs) (11-13 nm) capped with citric, ascorbic, and gallic acids], which are activated by high pulsed electric and electromagnetic fields allowing to overcome the nisin-resistance of bacteria. As a cell model the Gram-positive bacteria Bacillus subtilis and Gram-negative Escherichia coli were used. We have applied 10 and 30 kV cm-1 electric field pulses (100 µs × 8) separately and in combination with two pulsed magnetic field protocols: (1) high dB/dt 3.3 T × 50 and (2) 10 mT, 100 kHz, 2 min protocol to induce additional permeabilization and local magnetic hyperthermia. We have shown that the high dB/dt pulsed magnetic fields increase the antimicrobial efficiency of nisin NPs similar to electroporation or magnetic hyperthermia methods and a synergistic treatment is also possible. The results of our work are promising for the development of new methods for treatment of the drug-resistant foodborne pathogens to minimize the risks of invasive infections.