Attila Kántor

Identification of lactic acid bacteria isolated from wine using real-time PCR.

ABSTRACT Different lactic acid bacteria strains have been shown to cause wine spoilage, including the generation of substances undesirable for the health of wine consumers. The aim of this study was to investigate the occurrence of selected species of heterofermentative lactobacilli, specifically Lactobacillus brevis, Lactobacillus hilgardii, and Lactobacillus plantarum in six different Slovak red wines following the fermentation process. In order to identify the dominant Lactobacillus strain using quantitative (real time) polymerized chain reaction (qPCR) method, pure lyophilized bacterial cultures from the Czech Collection of Microorganisms were used. Six different red wine samples following malolactic fermentation were obtained from selected wineries. After collection, the samples were subjected to a classic plate dilution method for enumeration of lactobacilli cells. Real-time PCR was performed after DNA extraction from pure bacterial strains and wine samples. We used SYBR® Green master mix reagents for measuring the fluorescence in qPCR. The number of lactobacilli ranged from 3.60 to 5.02 log CFU mL−1. Specific lactobacilli strains were confirmed by qPCR in all wine samples. The number of lactobacilli ranged from 103 to 106 CFU mL−1. A melting curve with different melting temperatures (Tm) of DNA amplicons was obtained after PCR for the comparison of Tm of control and experimental portions, revealing that the most common species in wine samples was Lactobacillus plantarum with a Tm of 84.64°C.

Antibacterial Activity of Melissa officinalis L., Mentha piperita L., Origanum vulgare L. and Malva mauritiana against Bacterial Microflora Isolated from Fish

Abstract The aim of the present study was to detect the antibacterial activity of medicinal plants against fish microflora. A total of 4 ethanolic extracts of 6 plant species were collected from local environments of Slovakia and screened for antibacterial activity against bacterial microflora. Extracts of Melissa officinalis L., Mentha piperita L., Origanum vulgare L. and Malva mauritiana were used. Bacterial strains were isolated from common bleak (Alburnus alburnus) and common rudd (Scardinius erythrophthalmus) of Latvian origin. All bacterial strains were identified with the Matrix- Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS). Among fish microflora, Acinetobacter pittii, A. baumannii, Buttiauxella agrestis, Delftia acidovorans, Enterobacter cloacae, Serratia liquefaciens, Pseudomonas alcaligenes, Ps. oryzihabitans, Staphylococcus epidermidis, St. caprae, Pantoea agglomerans, Lelliottia amnigena, Providencia rettgeri, Escherichia coli and Rahnella aquatilis were identified. It has been shown that all plant extracts exhibit different degrees of antimicrobial activity against the tested bacteria. All bacterial strains in the present study were moderate sensitive to all extracts applied. The strongest antimicrobial effect of Malva mauritiana and Melissa officinalis L. against Pseudomonas oryzihabitans (6.67±1.53 resp. 9.67±0.58 mm) were found. The best antimicrobial activity of Mentha piperita L. was against Staphylococcus epidermis (7.33±0.58 mm) and strongest antimicrobial effect of Origanum vulgare L. was same against two bacterial strains Enterobacter cloacae and Serratia liquefaciens (9.67±0.58 mm).

Microbiological quality of chicken thighs meat after four essential oils combination, EDTA and vaccum packing

Normal 0 false false false CS JA X-NONE The aim of the present work to monitoring chicken the microbiological quality of vaccum packaged thighs after treatment by ethylenediaminetetraacetate (EDTA), anise ( Pimpinella anisum ), spearmint ( Mentha spicata var. crispa ), thyme ( Thymus vulgaris L.) oregano ( Origanum vulgare L.) essential oils and stored in at 4 ±0.5 °C for a period of 16 days. The following treatments of chicken thighs were used: air-packaged control samples, control vacuum-packaged samples, vacuum-packaging with EDTA solution 1.5% w/w, control samples, vacuum-packaging after treatment with Pimpinella anisum, Mentha spicata var. crispa essential oil at concentrations 0.2% v/w, vacuum-packaging after treatment with Thymus vulgaris L., Origanum vulgare L. essential oil at concentration 0.2% v/w. The quality assessment of all samples was done microbiologically and following microbiological parameters were detected: the anaerobic plate count, Enterobacteraceae counts, lactic acid bacteria and Pseudomonas spp. counts. The number of anaerobic plate count ranged from 3.69 log CFU.g -1 in all tested group on 0 day to 5.68 log CFU.g -1 on 16 day in control group stored in air condition. The number of lactic acid bacteria ranged from 2.00 log CFU.g -1 in all tested group on 0 day to 4.82 log CFU.g -1 on 16 day in group with oregano, thyme essential oils combination. Enterobacteriacea counts in chicken thighs was 0.68 log CFU.g -1 on 0 day to 7.58 CFU.g -1 on 16 day in air-packed meat samples . The Pseudomonas spp. was not found in all tested samples. Among the antimicrobial combination treatments examined in this work, the as application of vacuum packaging, EDTA and essential oils treatment was the most effective against the growth of Enterobactericeae , inhibitory effect on anaerobic plate count also was observed. The results of this present study suggest the possibility of application the Pimpinella anisum, Mentha spicata var. crispa , Thymus vulgaris L., Origanum vulgare L. essential oil of as natural food preservatives and potential sources of antimicrobial ingredients for food industry for chicken thighs meat treatment.

Application of lavender and rosemary essential oils improvement of the microbiological quality of chicken quarters

The aim of the present work was monitoring of chicken quarters microbiological indicators after treatment by ethylenediaminetetraacetate (EDTA), lavender ( Lavandula angustifolia L.) and rosemary ( Rosmarinus officinalis L.) essential oil, stored under vacuum packaging, at 4 ±0.5°C for a period of 16 days. The following treatments of chicken quarters were used: Air-packaging control samples, control vacuum-packaging samples, vacuum-packaging with EDTA solution 1.50% w/w, control samples, vacuum-packaging with Lavandula angustifolia essential oil at concentrations 0.2% v/w and vacuum-packaging with Rosmarinus officinalis essential oil at concentration 0.2% v/w. The quality assessment of all samples was established by microbiological analysis. Sampling was carried out after certain time intervals: 0, 4, 8, 12 and 16 days. Chicken quarters were stored under vacuum packaging, at 4 ±0.5°C during experiment. Microbiological analyses were conducted by using standard microbiological methods. Anaerobic plate count were determined using Plate Count Agar, after incubation for 2 days at 35°C under anaerobic condition. Pseudomonas spp. were determined on Pseudomonas Isolation agar after incubation at 48 h at 25°C. For lactic acid bacteria were inoculated into Rogosa and Sharpe agar after incubation 48-78 h at 37°C in an aerobic atmosphere supplemented with carbon dioxide (5% CO 2 ). For members of the family Enterobacteriaceae violet red bile glucose agar were used and samples were incubated at 37°C for 24 h. The initial APC value of chicken quarter was 3.00 log CFU.g -1 on 0 day. The number of anaerobic plate count ranged from 3.00 log CFU.g -1 in all tested group on 0 day to 6.11 log CFU.g -1 on 16 day in control group stored in air condition. The initial LAC value of chicken quarter was 3.00 log CFU.g -1 on 0 day. The number of lactic acid bacteria ranged from 3.00 log CFU.g -1 in all tested group on 0 day to 3.58 log CFU.g -1 on 16 day in control group stored in air condition. The initial Enterobacteriacea genera value of chicken quarter was 2.00 log CFU.g -1 on 0 day. Presences of these bacteria were found on all groups at 16 days. The results of this present study suggest the possibility of application the Lavandula angustifolia and Rosmarinus officinalis essential oil as natural food preservatives and potential sources of antimicrobial ingredients for food industry.

Determination of Biological and Sensory Profiles of Biscuits Enriched with Tea (Camellia sinensis L.) Powder

Abstract The aim of this work was to characterise the biological and sensory profile of biscuits enriched with green (1 and 3%) and black tea (1 and 3%) powders. Biscuits without the addition of tea were used as a control. Phenolic concentration, flavonoid concentration, and antioxidant activity were determined spectrophotometrically. Amino acid composition was determined using automatic amino acid analyser AAA 400 and crude fibre content using an Ancom analyser. Sensory profiles were evaluated by comparison of enriched and control biscuit samples. The enriched biscuits showed higher phenolic and flavonoid concentration and antioxidant activity estimated by DPPH and phospholybdenum method in comparison with levels in the control group. The best results for antioxidant activity estimated by DPPH and phosphomolybdenum methods were achieved in biscuits enriched with black tea powder (3%): 2.25 and of 32.64 mg TEAC·g−1, respectively. Total phenolic concentration was 1.16 mg GAE·g−1, and total flavonoid concentration was 0.13 mg QE·g−1. These biscuits had higher concentration of crude fibre in comparison with the control group and the highest concentration (0.64%) was found in biscuits with addition of 3% green tea powder. The amino acid composition in samples, including in the control sample was balanced, with slightly higher concentration of threonine, serine, and methionine in enriched samples, but this parameter was not statistically significant. Biscuits enriched with green and black tea had higher sensory scores for taste, smell and aftertaste.

Bacteria and yeasts isolated from different grape varieties

Patasiova Martina 14.00 The aim of this study was to isolate and identify bacteria and yeasts in different grape samples. The samples were collected in September 2017. Used 13 grape samples in this study (9 white and 4 red) were from the local Slovak winemakers. Alibernet, Irsai Oliver, Dornfelder, Blue Frankish, Feteasca regala, Green Veltliner, Palava, Mūller Thurgau, Rhinriesling, Cabernet Savignon, Pinot Blanc, Savignon Blanc and Welschriesling. Two cultivation media were used for detection of bacteri and yeasts in grape samples. Malt extract agar base (MEA) and Tryptone Soay agar (TSA) were used for the cultivation of bacteria and yeasts. Cultivation was performed by spread plate method. Ethanol/formic acid extraction procedure was used for preparation of samples. MALDI-TOF Mass Spectrometer (Microflex LT/SH) (Bruker Daltonics, Germany) was used for identification of bacteria and yeasts. In total, 8 genera of yeasts, 8 genera of Gram-negative bacteria and 10 of Gram-positive bacteria were identified. Together 333 isolates, yeasts, Gram-negative and Gram-positive bacteria were identified. Normal 0 21 false false false EN-US X-NONE X-NONE

Microscopic fungi isolated from different Slovak grape varieties

Normal 0 21 false false false EN-GB X-NONE X-NONE The aim of this study was to isolate and identify microscopic fungi in different grape samples. We collected 13 grapes varienties samples (9 white and 4 red) from local Slovak winemakers in the end of the September 2017. Used 13 grape samples in this study: Alibernet, Irsai Oliver, Dornfelder, Blue Frankish, Feteasca regala, Green Veltliner, Palava, Mūller Thurgau, Rhinriesling, Cabernet Savignon, Pinot Blanc, Savignon Blanc and Welschriesling. Microscopic fungi in grape samples were detected on Malt extract agar by spread plate method. The number of microscopic fungi ranged from 2.85 log cfu.g -1 in Cabernet Savignon to 4.83 log cfu.g -1 in Feteasca regala. A total of 627 isolates of microscopic fungi were obtained in this study. The most abundant fungi belonged to genera Alternaria and Penicillium (100% frequency). The high frequency was also detected for Aspergillus (76.92%) and Cladosporium (76.92%) but with lesser relative density. Alternaria sp., Aspergillus niger , Aspergillus sp., Botrytis cinerea , Cladosporium sp., Penicillium expansum , Phoma sp., Rhizopus sp. and Trichoderma sp. species were isolated from grape berries.

IN VITRO EFFECTS OF ACINETOBACTER BAUMANNII AND SELECTED NATURAL BIOMOLECULES ON RABBIT SPERMATOZOA MOTILITY

The aim of this study was to assess the potential efficiency of selected biologically active substances on the motility behaviour of rabbit spermatozoa subjected to in vitro induced A. baumannii contamination. The semen samples used for A. baumannii detection were collected from 10 New Zealand white male rabbits and the presence of the bacterium was confirmed using MALDI-TOF Mass Spectrometry. For the in vitro experiments rabbit spermatozoa were re-suspended in PBS, containing mineral supplements, BSA and glucose in the presence of 3x105 CFU A. baumannii and diverse concentrations of selected biomolecules (resveratrol - RES, quercetin - QUE, curcumin - CUR, epicatechin - EPI, isoquercitrin - ISO). The sperm motility was assessed using the computer-aided sperm analysis at 0h, 2h, 4h and 6h. A. baumannii significantly decreased the sperm motility (P<0.001) at Time 2h and maintained this negative impact throughout the in vitro culture. Meanwhile, the motility at Time 2h was significantly higher in the samples subjected to A. baumannii together with 10 μmol/L RES (P<0.01); 5, 10 and 50 μmol/L QUE (P<0.001); 1 μmol/L CUR (P<0.05); 10, 50 and 100 μmol/L EPI (P<0.01) as well as 50 μmol/L (P<0.05) and 100 μmol/L ISO (P<0.001) in comparison to the control exposed to the bacterium exclusively. After 4h, the motility remained significantly higher in the groups co-treated with the inoculum and 10 μmol/L RES (P<0.05), 50 μmol/L QUE (P<0.05) as well as 50 μmol/L EPI (P<0.05) when compared to the positive control. Nevertheless, none of the biomolecules was effective against the rapid decline of sperm motility caused by A. baumannii during later stages of the experiment (Time 6h). Based on these results, one can conclude that RES, QUE and EPI exhibit antibacterial properties providing a selective advantage to spermatozoa in the presence of A. baumannii, particularly during short-term rabbit semen handling.

Antimicrobial Effect of Sage (Salvia officinalis L.) and Rosemary (Rosmarinus officinalis L.) Essential Oils on Microbiota of Chicken Breast

Abstract The aim of the study was to evaluate the antimicrobial effect of sage and rosemary essential oils (EO) on microbiota of fresh chicken breast. Sample treatments were stored without packaging, vacuum-packaged, vacuum-packaged with EDTA 1.5% v/w, sage and rosemary EO treatment 0.2% v/w. Assessment of food quality was done by anaerobic plate count (APC), and Enterobacteriaceae, lactic acid bacteria (LAB) and Pseudomonas spp. counts a period of 16 days of storage at 4 ± 0.5 °C. Bacterial species were identified with a MALDI TOF MS Biotyper. Antimicrobial activity of isolates against both EO were tested. The APC varied from 2.97 log CFU/g to 6.81 log CFU/g, LAB from 2.35 log CFU/g to 3.36 log CFU/g and Enterobacteriaceae from 0.00 log CFU/g on day 0 to 4.77 log CFU/g with the highest counts on day 16 and in control unpackaged samples. Pseudomonas spp. was found only on days 0, 4, 8, and 12, with counts from 0.00 log CFU/g on day 16 to 2.89 log CFU/g on day 4 in control unpackaged samples. APC were represented by Staphylococcus and Kocuria, LAB with Lactobacillus and Enterobacteriaceae with Buttiauxella, Escherichia, Hafnia, Serratia and Yersinia. The Pseudomonas genus was represented by ten species. The best antimicrobial effect on APC, Enterobacteriaceae, LAB and Pseudomonas was achieved by application of EO. The results suggest the potential use of Salvia officinalis L. and Rosmarinus officinalis L. EOs as natural food preservatives and potential sources of antimicrobial ingredients in the food industry.

Microorganisms of Grape Berries

Abstract Grape surface is an unstable habitat that changes greatly according to the stage of grape ripening. Different bacteria and yeasts can colonise the surface of grape berry and the diversity of microorganisms depends on the stage of ripening, pesticide application and health condition. The aim of this study was to study the microflora of the surface of grape berries. Altogether, 19 grape samples from Slovakia were collected. The spread plate method was applied and a 100 μL inoculum of each dilution (10−2, 10−3) was plated on TSA, MEA, and MRS agar for isolation of microorganisms from grapes. Proteins were extracted from cells by ethanol/formic acid extraction procedure. MALDI-TOF Mass Spectrometry was used for identification of microorganisms. In total, 11 genera of Gram-negative bacteria, 11 of Gram-positive bacteria and nine of yeasts were identified. Among 200 isolates, Gram-negative, Gram-positive bacteria and yeasts represented 11%, 27% and 62% of the total number of isolates studied. The most common genera of isolated yeasts were Hanseniaspora (37%), Metschnikowia (31%), and Rhodotorula (10%). The most frequently isolated among Gram-negative bacteria were Acinetobacter (22%), Pseudomonas (22%) and Sphingomonas (13%). The most common genera of Gram-positive bacteria were Bacillus (20%), Lactobacillus (19%), Leuconostoc and Staphylococcus (11%), respectively.