The aim of the present work was to study the mode of the induction of the biosynthesis of macedocin, the lantibiotic produced by Streptococcus macedonicus ACA-DC 198. Macedocin was produced when the ...strain was grown in milk but not in MRS or M17 broth. No autoinduction mechanism was observed. Production did not depend on the presence of lactose or galactose in the culture medium or on a coculture of the producer strain with macedocin-sensitive or macedocin-resistant strains. Induction seemed to depend on the presence of one or more heat-stable protein components produced when S. macedonicus ACA-DC 198 was grown in milk. The partial purification of the induction factor was performed by a combination of chromatography methods, and its activity was confirmed by a reverse transcription-PCR approach (RT-PCR). Mass spectrometric (MS) and tandem mass spectrometric (MS/MS) analyses of an induction-active fraction showed the presence of several peptides of low molecular mass corresponding to fragments of αS₁- and β-casein as well as β-lactoglobulin. The chemically synthesized αS₁-casein fragment 37-55 (2,253.65 Da) was proven to be able to induce macedocin biosynthesis. This is the first time that milk protein degradation fragments are reported to exhibit a bacteriocin induction activity.
Goat milk yogurt (GMY) and raisins are popular foods with a favourable nutrient profile. Our aim was to determine the glycaemic index (GI) and postprandial responses to GMY-containing ...angiotensin-converting enzyme inhibitory (ACE-I) peptides carrying the RPKHPINHQ isracidin fragment and two Greek raisin varieties in an acute feeding setting. A total of twelve healthy participants (four male and eight female) consumed breakfast study foods containing 25 g available carbohydrate on seven occasions over a 3- to 9-week period: food 1: D-glucose (25 g) served as the control and was consumed on three separate occasions; food 2: GMY (617·28 g); food 3: Corinthian raisins (37·76 g); food 4: Sultana raisins (37·48 g) and food 5: GMY & C (308·64 g GMY and 18·88 g C). Postprandial glucose was measured over a 2 h period for the determination of GI and glycaemic load (GL). Subjective appetite ratings (hunger, fullness and desire to eat) were assessed by visual analogue scales (100 mm) at 0–120 min. Blood pressure (systolic and diastolic; BP) was measured at baseline and 120 min. GMY provided low GI (26), C and S provided high GI/low GL (75/10 and 70/9, respectively) and GMYC provided low GI (47) values on glucose scale compared with D-glucose. Peak blood glucose rise was significantly lower only for GMY and GMYC compared with reference food (D-Glucose), as well as C and S (P
for all < 0·05). No differences were observed between test foods for fasting glucose, BP and subjective appetite. In conclusion, GMY and GMYC attenuated postprandial glycaemic responses, which may offer advantages to glycaemic control.
Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS), a culture based alternative for microbial diversity studies, is an attractive tool to dereplicate large ...numbers of isolates to a smaller set of representatives for downstream characterization. In the present study, MALDI-TOF MS, combined with a database of reference spectra compiled in previous studies, was applied to identify 88 non-starter lactic acid bacteria (NSLAB) isolated from 18 samples of four different artisanal cheeses produced in the Island of Naxos, Greece, from raw sheep and goat milk without the addition of starters. Eighty-four isolates (95.5%) could be identified directly via MALDI-TOF MS. Lactobacillus brevis and Lactobacillus plantarum were the dominant species, followed by Lactococcus lactis, Leuconostoc mesenteroides Lactobacillus paracasei, Lactobacillus rhamnosus, Pediococcus pentosaceus and Enterococcus faecium. The remaining four isolates represented species present in the database; however, within-species diversity was insufficiently covered. Additionally, pheS sequencing was applied to confirm identification.
•Application of MALDI-TOF MS for the identification of lactic acid bacteria•First fingerprinting of the non-starter lactic acid bacteria of artisanal cheeses produced in the Island of Naxos, Greece•PheS sequencing was performed to validate MALDI-TOF MS identication
Artisanal cheeses, in particular those prepared from raw milk, are the most commonly used ecosystems to mine microorganisms with both technological and human-health related potential. In the present ...study, the microbiota of two Greek artisanal cheeses, the PDO Kalathaki and the non-PDO Melichloro, was explored via classical microbiological analysis. Lactic acid bacteria isolated mainly belonged to the genus Enterococcus (66%) and to a lesser degree to Pediococcus (15%), Leuconostoc (8%), Lactobacillus (6%) and Lactococcus (5%). From the 112 initial isolates, 32 were selected and studied for their probiotic potential, i.e. safety traits, survival under gastrointestinal tract conditions, antimicrobial activity, adhesion, angiotensin I-converting enzyme inhibitory activity and immunomodulation of human monocytes. Two isolates, namely Lactobacillus brevis ACA-DC 1705 and Leuconostoc mesenteroides ACA-DC 1738, exhibited the most promising probiotic potential due to strong angiotensin I-converting enzyme inhibitory activity and anti-inflammatory modulation of immune cells, respectively, and, thus they could be used as adjuncts in novel functional products.
•First report on the microbiota diversity of the cheeses Kalathaki and Melichloro.•Evaluation of the probiotic potential of the isolated lactic acid bacteria.•A Lactobacillus brevis showed angiotensin I-converting enzyme inhibitory activity.•A Leuconostoc mesenteroides displayed anti-inflammatory modulation of immune cells.
Staka is a naturally fermented cream produced from sheep or mixtures of sheep and goat milk in the island of Crete, Greece. This work is the first report on Staka cream microbiota. Initially, counts ...of various bacterial groups were enumerated using selective media and growth conditions. All isolates (101) were subjected to matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) analysis; the spectra obtained were compared with those of an in-house reference database. While 70 isolates were considered identified at the species level, the remaining 31 were not, probably due to a lack of, or an insufficient number of, reference spectra in the MALDI-TOF MS database. Strains representative of the observed diversity among the unidentified MALDI-TOF MS spectra were therefore subjected to pheS and 16S rRNA gene sequencing analysis for further identification. The identification results revealed the presence of Lactobacillus spp., Enterococcus spp., Streptococcus thermophilus and Aerococcus viridans.
•Microemulsions based on four essential oils were used to encapsulate nisin.•Structural changes were revealed regarding the micellar interface flexibility.•Nisin loaded systems showed antimicrobial ...efficacy against pathogens.•Dittany-based microemulsions had the best antimicrobial efficacy against pathogens.•The system’s antimicrobial efficacy depended on microemulsions’ characteristics.
During the last years, the food industry is working on the replacement of high energy methodologies with more sustainable techniques for the encapsulation of natural preservatives, in order to enhance their effectiveness as food additives. In the present study, nisin, an antimicrobial agent, was encapsulated in essential oil-containing microemulsions. More specifically, rosemary, thyme, oregano, and dittany essential oil-containing microemulsions were formulated to encapsulate nisin enhancing the system’s overall antimicrobial activity. The systems were investigated for the interfacial properties and size of the surfactants’ monolayer using electron paramagnetic resonance spectroscopy and dynamic light scattering. Subsequently, nisin-loaded microemulsions were tested for their antimicrobial activity against Lactococcus lactis, Staphylococcus aureus, Listeria monocytogenes, and Bacillus cereus, using the well diffusion assay. Finally, this technique was validated by a killing assay. Overall, this study provides important information on the antibacterial activity of nisin-loaded nano-carriers enhanced by essential oils, in relation to the microemulsions’ structure.
•Reverse micelles in vegetable oils were used as nanocarriers to encapsulate nisin.•Structural system characterization revealed some changes in the presence of nisin.•The loaded systems showed ...antimicrobial efficacy against pathogens in real foods.•The nanocarriers’ antimicrobial efficacy depended on food matrices’ composition.•Ethanol may play an important role on the interactions with food matrices.
Reverse micelles (RMs) as nanocarriers of nisin were optimized for the highest water and bacteriocin content. RMs formulated with either refined olive oil or sunflower oil, distilled monoglycerides, ethanol, and water were effectively designed. Structural characterization of the RMs was assessed using Electron Paramagnetic Resonance Spectroscopy and Small Angle X-ray Scattering in the presence and absence of nisin. No conformational changes occurred in the presence of nisin for the nanocarriers. To assess efficacy of the loaded systems, their antimicrobial activity against Staphylococcus aureus and Listeria monocytogenes was tested in lettuce leaves and minced meat, respectively. Antimicrobial activity was evident in both cases. Interestingly, a synergistic antimicrobial effect was observed in lettuce leaves and to a lesser extent in minced meat between nisin and some of the nanocarriers’ constituents (probably ethanol). Our findings suggest complex interactions that take place when RMs are applied in different food matrices.
The technological and probiotic potential of lactic acid bacteria isolated from artisanal Greek yoghurt and fermented milks were evaluated. Fifty-three strains were identified by rep-PCR and 16S rDNA ...sequencing to belong to different Lactobacillus or Enterococcus spp., as well as to Streptococcus thermophilus and Lactococcus lactis. Several strains exhibited promising technological and probiotic properties. Among them, we focused on the production of bioactive peptides with angiotensin-converting enzyme inhibitory (ACE-I) activity during milk fermentation. The majority of strains produced ACE-I peptides when grown in skimmed milk. ACE-I peptides were sometimes sequestered in the original fermented milk sample, but were released and detected following high performance liquid chromatography (HPLC) purification. Mass spectrometry analysis of major peptide peaks in HPLC fractions with ACE-I activity revealed that they derived from the N- or C-terminal of the isracidin peptide region of αS1-casein and two internal peptide fragments, one from β-casein and one from κ-casein.
Probiotic Feta cheese was produced using Propionibacterium freudenreichii subsp. shermanii as an adjunct since it has been shown to induce apoptosis of colon cancer cells in vitro and in vivo through ...the production of propionate and acetate. Microbiological and physicochemical analysis of the cheese was performed through the 60 d period of ripening. Counts of propionibacteria increased until day 7 and then remained constant until the end of ripening (approximately 9 log cfu g−1). Moreover, throughout ripening the presence of P. freudenreichii subsp. shermanii LMG 16424T was confirmed by 16S rRNA gene sequence analysis, while propionic acid was first detected on day 7 and reached a concentration of 52.1 mm on day 60. The ripened cheese containing P. freudenreichii subsp. shermanii was very well accepted by the sensory evaluation panellists. This is the first time that P. freudenreichii subsp. shermanii was studied as a probiotic adjunct in a white-brined cheese.
Lactobacillus curvatus L442, isolated from Greek traditional fermented sausage prepared without the addition of starters, produces a bacteriocin, curvaticin L442, which is active against the pathogen ...Listeria monocytogenes. The bacteriocin was purified by 50% ammonium sulphate precipitation, cation exchange, reverse phase and gel filtration chromatography. Partial N-terminal sequence analysis using Edman degradation revealed 30 amino acid residues, revealing high homology with the amino acid sequence of sakacin P. Curvaticin L442 is active at pH values between 4.0 and 9.0 and it retains activity even after incubation for 5 min at 121 degrees C with 1 atm of overpressure. Proteolytic enzymes and alpha-amylase inactivated this curvaticin, while the effect of lipase was not severe.
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