Safety and Beneficial Properties of Bacteriocinogenic Lactococcus lactis and Pediococcus pentosaceus Strains, and Their Effect Versus Oral Cavity Related and Antibiotic-Resistant Pathogens.

Pediococcus pentosaceus 732, Lactococcus lactis subsp. lactis 431, and Lactococcus lactis 808, bacteriocinogenic strains previously isolated from kimchi and banana, were investigated for their safety, beneficial properties and in vitro inhibition of pathogens such as Listeria monocytogenes ATCC 15313 and Staphylococcus simulans KACC 13241 and Staphylococcus auricularis KACC 13252. The results of performed physiological, biochemical, and biomolecular tests suggest that these strains can be deemed safe, as no virulence genes were detected in their DNA. Notably, only the gad gene associated with GABA production was identified in the DNA isolated of Lc. lactis 808 and Lc. lactis subsp. lactis 431 strains. All tested LAB strains exhibited γ-hemolysins and were non-producers of gelatinase and biogenic amines, which suggested their safety potential. Additionally, they were relatively susceptible to antibiotics except for streptomycin, tobramycin, and vancomycin for Pd. pentosaceus 732. The growth of Pd. pentosaceus 732, Lc. lactis subsp. lactis 431, and Lc. lactis 808 and their survival were minimally affected by up to 3% ox bile and low pH (except pH 2.0 and 4.0). Moreover, these LAB strains were not inhibited by various commercial extracts as well as most of the tested medications tested in the study. They did not produce proteolytic enzymes but exhibited production of D/L-lactic acid and ß-galactosidase. They were also hydrophilic. Furthermore, their survival in artificial saliva, gastric simulation, and enteric passage was measured followed by a challenge test to assess their ability to inhibit the selected oral pathogens in an oral saliva model conditions.

Antimicrobial Peptides (Bacteriocins) Produced by Lactococcus lactis and Pediococcus pentosaceus Strains with Activity Against Clinical and Food-Borne Pathogens.

Bacteriocins are ribosomal-synthesized peptides with antimicrobial activity, produced by different groups of bacteria, including lactic acid bacteria (LAB). Most of the produced by LAB bacteriocins can be described with rather broad spectra of inhibition and they offer suggested applications in food preservation and pharmaceutical sector. Different LAB were isolated from fermented food products and fruits, obtained from the region of Pohang, Korea, and identified based on physiological, biochemical, and molecular methods. The promising isolates, Pediococcus pentosaceus 732, Lactococcus lactis 808, and Lactococcus lactis subsp. lactis 431, were identified based on biochemical, physiological, and biomolecular approaches, including 16S rRNA partial sequencing, and were evaluated for production of bacteriocin, including stability in presence of enzymes, chemicals, pH, and temperatures. Adherence properties for the expressed bacteriocins by P. pentosaceus 732, Lc. lactis 808, and Lc. lactis subsp. lactis 431 were evaluated at presence of selected chemicals, pH, and temperatures. The presence of bacteriocin genes in the strains was investigated and analyzed. The bacterial effect of bacteriocin produced by studied strains on Listeria spp. and Staphylococcus spp. has been shown for actively growing and stationary cells. Similar growth and bacteriocin production were observed when studied strains were cultured in MRS at 30 °C or 37 °C. The presence of nisin operon with some point mutations on the genomic DNA was recorded based on the performed PCR reactions targeting different genes associated with nisin expression for both lactococcal strains. Pediocin PA-1 operon was evaluated in a similar manner for P. pentosaceus 732.

Bacillus Strains Isolated from Korean Fermented Food Products with Antimicrobial Activity Against Staphylococci, an Alternative for Control of Antibiotic-Resistant Pathogens.

Multidrug resistance in foodborne and clinical pathogens is a worldwide health problem. The urgent need for new alternatives to the existing antibiotics is emerging. Bacteriocin-like inhibitory substances can be considered part of the new generation of antimicrobials, which can be potentially applied in the food industry and health care practices. This study aimed to select Bacillus strains with antimicrobial activity against Staphylococcus spp. with future application in the formulation of pharmaceutical antimicrobial preparations. Putative antimicrobial agent-producing strains, previously isolated and preidentified as Bacillus spp. were profiled by repetitive element sequence-based polymerase chain reaction (rep-PCR) and 16s rRNA sequencing identified the strains as Bacillus tequilensis ST1962CD with 99.47% identity confidence and as Bacillus subtilis subsp. stercoris ST2056CD with 98.45% identity confidence. Both the selected Bacillus strains were evaluated via biomolecular and physiological approaches related to their safety and virulence, beneficial properties, enzyme production profile, and presence of corresponding genes for the production of antimicrobials and virulence. Both strains were confirmed to harbor srfa and sbo genes and be free of hemolysin binding component (B) and two lytic components (L1 and L2) [BL] and nonhemolytic enterotoxin-associated genes. Produced antimicrobial agents by strains ST1962CD and ST2056CD were partially purified through the combination of ammonium sulfate precipitation and hydrophobic-based chromatography on SepPakC18 and evaluated regarding their cytotoxicity. The dynamics of bacterial growth, pH change, accumulation of produced antimicrobials, and the mode of action were evaluated. Obtained results were pointing to the potential application of safe B. tequilensis ST1962CD and B. subtilis subsp. stercoris ST2056CD strains as functional beneficial microbial cultures that are putative producers of surfactin and/or subtilosin, as potent antimicrobials, for the treatment of some staphylococcal-associated infections. Expressed antimicrobials were shown to be not cytotoxic, and appropriate biotechnological approaches need to be developed for cost-effective production, isolation, and purification of expressed antimicrobials by studied strains.

The Evaluation of Different Bacteriocinogenic Bacillus spp. with Activity Against Staphylococcus spp. and Their Beneficial and/or Hazardous Properties.

The aim of this project was to screen for bacteriocinogenic Bacillus strains with activity versus Staphylococcus spp. with future application in formulation of pharmaceutical antimicrobial preparations. Putative bacteriocinogenic strains, isolated and pre-identified as Bacillus spp. were selected for future study and differentiated based on repPCR and identified as Bacillus subtilis for strains ST826CD and ST829CD, Bacillus subtilis subsp. stercoris for strain ST794CD, Bacillus subtilis subsp. spizizenii for strain ST824CD, Bacillus velezensis for strain ST796CD, and Bacillus tequilensis for strain ST790CD. Selected strains were evaluated regarding their safety/virulence, beneficial properties, and potential production of antimicrobials based on biomolecular and physiological approves. Expressed bacteriocins were characterized regarding their proteinaceous nature, stability at different levels of pH, temperatures, and the presence of common chemicals applied in bacterial cultivation and bacteriocin purification. Dynamic of bacterial growth, acidification, and cumulation of produced bacteriocins and some aspects of the bacteriocins mode of action were evaluated. Based on obtained results, isolation and application of expressed antimicrobials can be realistic scenario for treatment of some staphylococcal associated infections. Appropriate biotechnological approaches need to be developed for cost effective production, isolation, and purification of expressed antimicrobials by studied Bacillus strains.

Beneficial features of pediococcus: from starter cultures and inhibitory activities to probiotic benefits.

Pediococci are lactic acid bacteria (LAB) which have been used for centuries in the production of traditional fermented foods. There fermentative abilities were explored by the modern food processing industry in use of pediococci as starter cultures, enabling the production of fermented foods with distinct characteristics. Furthermore, some pediococci strains can produce bacteriocins and other antimicrobial metabolites (AMM), such as pediocins, which are increasingly being explored as bio-preservatives in various food matrices. Due to their versatility and inhibitory spectrum, pediococci bacteriocins and AMM are being extensively researched not only in the food industry, but also in veterinary and human medicine. Some of the pediococci were evaluated as potential probiotics with different beneficial areas of application associated with human and other animals' health. The main taxonomic characteristics of pediococci species are presented here, as well as and their potential roles and applications as starter cultures, as bio-preservatives and as probiotic candidates.

Characterization and safety evaluation of two beneficial, enterocin-producing Enterococcus faecium strains isolated from kimchi, a Korean fermented cabbage.

Enterococcus faecium ST20Kc and ST41Kc were isolated from kimchi, a traditional Korean fermented cabbage. Bacteriocins produced by both strains exhibited strong activity against Listeria monocytogenes and various Enterococcus spp., including 30 vancomycin-resistant enterococcal strains, but not against other lactic acid bacteria (LAB) on the evaluated test panel. The antimicrobials produced by the strains were found to be proteinaceous and stable even after exposure to varying pH, temperature, and chemicals used in the industry and laboratory processes. Antimicrobial activity of both strains was evaluated as bactericidal against exponentially growing cultures of L. monocytogenes ATCC® 15313™ and Enterococcus faecalis 200A. Based on tricine-SDS-PAGE, the molecular weights of the bacteriocins produced by the strains were between 4 and 6 kDa. Additionally, both strains were susceptible to antibiotics, including vancomycin, kanamycin, gentamycin, ampicillin, streptomycin, tylosin, chloramphenicol, clindamycin, and tetracycline. Adhesion genes, map, mub, and EF-Tu, were also detected in the genomes of both strains. With gastrointestinal stress induction, both strains showed high individual survival rates, and capability to reduce viable counts of L. monocytogenes ATCC® 15313™ and Enterococcus faecalis 200A in mixed cultures. Based on the metabolomics analysis, both strains were found to produce additional antimicrobial compounds, particularly, lactic acid, phenyllactic acid, and phenethylamine, which can be potentially involved in the antimicrobial interaction with pathogenic microorganisms.

Do Your Kids Get What You Paid for? Evaluation of Commercially Available Probiotic Products Intended for Children in the Republic of the Philippines and the Republic of Korea.

A wide range of probiotic products is available on the market and can be easily purchased over the counter and unlike pharmaceutical drugs, their commercial distribution is not strictly regulated. In this study, ten probiotic preparations commercially available for children's consumption in the Republic of the Philippines (PH) and the Republic of Korea (SK) have been investigated. The analyses included determination of viable counts and taxonomic identification of the bacterial species present in each formulation. The status of each product was assessed by comparing the results with information and claims provided on the label. In addition to their molecular identification, safety assessment of the isolated strains was conducted by testing for hemolysis, biogenic amine production and antibiotic resistance. One out of the ten products contained lower viable numbers of recovered microorganisms than claimed on the label. Enterococcus strains, although not mentioned on the label, were isolated from four products. Some of these isolates produced biogenic amines and were resistant to one or several antibiotics. Metagenomic analyses of two products revealed that one product did not contain most of the microorganisms declared in its specification. The study demonstrated that some commercial probiotic products for children did not match their label claims. Infants and young children belong to the most vulnerable members of society, and food supplements including probiotics destined for this consumer group require careful checking and strict regulation before commercial distribution.