Miroslava Kačániová

Critical review on biofilm methods

Abstract Biofilms are widespread in nature and constitute an important strategy implemented by microorganisms to survive in sometimes harsh environmental conditions. They can be beneficial or have a negative impact particularly when formed in industrial settings or on medical devices. As such, research into the formation and elimination of biofilms is important for many disciplines. Several new methodologies have been recently developed for, or adapted to, biofilm studies that have contributed to deeper knowledge on biofilm physiology, structure and composition. In this review, traditional and cutting-edge methods to study biofilm biomass, viability, structure, composition and physiology are addressed. Moreover, as there is a lack of consensus among the diversity of techniques used to grow and study biofilms. This review intends to remedy this, by giving a critical perspective, highlighting the advantages and limitations of several methods. Accordingly, this review aims at helping scientists in finding the most appropriate and up-to-date methods to study their biofilms.

Intra- and inter-species interactions within biofilms of important foodborne bacterial pathogens.

A community-based sessile life style is the normal mode of growth and survival for many bacterial species. Under such conditions, cell-to-cell interactions are inevitable and ultimately lead to the establishment of dense, complex and highly structured biofilm populations encapsulated in a self-produced extracellular matrix and capable of coordinated and collective behavior. Remarkably, in food processing environments, a variety of different bacteria may attach to surfaces, survive, grow, and form biofilms. Salmonella enterica, Listeria monocytogenes, Escherichia coli, and Staphylococcus aureus are important bacterial pathogens commonly implicated in outbreaks of foodborne diseases, while all are known to be able to create biofilms on both abiotic and biotic surfaces. Particularly challenging is the attempt to understand the complexity of inter-bacterial interactions that can be encountered in such unwanted consortia, such as competitive and cooperative ones, together with their impact on the final outcome of these communities (e.g., maturation, physiology, antimicrobial resistance, virulence, dispersal). In this review, up-to-date data on both the intra- and inter-species interactions encountered in biofilms of these pathogens are presented. A better understanding of these interactions, both at molecular and biophysical levels, could lead to novel intervention strategies for controlling pathogenic biofilm formation in food processing environments and thus improve food safety.

Antioxidant and antimicrobial properties of monofloral bee pollen

The main aim of this study was to determine antioxidant properties and antibacterial activity of monofloral bee pollen samples to pathogenic bacteria. These samples were collected in different localities in Slovakia. The antioxidant properties of examined plant species were different and decreasing in the following order: Brassica napus subsp. napus L > Papaver somniferum L. > Helianthus annuus L. The antimicrobial effect of the bee product samples were tested by using the agar well diffusion method. The methanol (99.9% and 70%) and the ethanol (96% and 70%) were used for extraction. In this study, five different strains of bacteria were tested: Listeria monocytogenes CCM 4699; Pseudomonas aeruginosa CCM 1960; Staphylococcus aureus CCM 3953; Salmonella enterica CCM 4420; and Escherichia coli CCM 3988. The most sensitive bacteria of the poppy pollen ethanolic extract was Staphylococcus aureus was (70%) The most sensitive bacteria of rape bee pollen methanolic extract (70%) and sunflower ethanolic extract (70%) was Salmonella enterica.

The effect of selected microbial strains on internal milieu of broiler chickens after peroral administration.

The aim of this work was to investigate the effect of probiotic preparation with multi-strains composition (Lactobacillus delbrueckii ssp. bulgaricus LAT 187, Lactobacillus acidophilus LAT 180, Lactobacillus helveticus LAT 179, Lactobacillus delbrueckii ssp. lactis LAT 182, Streptococcus thermophilus LAT 205 and Enterococcus faecium E 253) on internal milieu, antioxidant status and body weight of broiler chickens. Chickens were divided into three groups (C, control; E1 and E2, experimental groups). Experimental chickens received the probiotic preparation in drinking water. Probiotic preparation caused a significant increase of serum calcium and potassium content. In the group with higher dose of probiotic strains serum triglycerides level decreased. Total antioxidant status in groups with addition of probiotic strains showed higher values in comparison to control group. Serum albumin level was found to increase after consumption of probiotic preparation in group with higher dose of probiotic strains. Probiotic strains improved body weight in last observed weeks of feeding.

Environmental concentration of selected elements and relation to physicochemical parameters in honey

The purpose of this study was to evaluate the content of selected elements and physicochemical indicators of the honey of Apis mellifera. Samples of blossom, blend and honeydew honeys were randomly obtained from the Eastern, Western and Central of Slovakia. Contents of elements were measured with the use of flame atomic absorption spectrophotometry (FAAS) method and anodic stripping voltammetry (ASV) method with CGMDE as working electrode. The content of heavy metals (mg/kg dry weight) ranged as follows: Zn 0.25–3.82; Cu 0.12–1.37; Ni 0.06–0.60; Mg 0.10–50.43; Ca 10.32–87.21; Pb and Cd were under detectable limit (0.001 mg/kg). The physical and chemical indicators ranged as follows: moisture (%) 16.00–19.80; HMF (mg/kg) 10.00–40.00; pH 3.58–5.12; conductivity (mS/m) 13.45–99.12 and a w 0.456–0.659. Significant difference (P < 0.05) for elements and physicochemical indicators was analyzed. Significant correlations between Cu – Zn, Cu – Ni, Cu – Mg, Ca – Zn and Ca – Mg were detected. With reference to Codex Alimentarius of Slovakia, maximal levels of heavy metals content and selected physicochemical properties were not exceeded. On average, the content of trace elements decreased in the order: Ca > Mg > Zn > Cu > Ni and physicochemical indicators decreased: conductivity > HMF > moisture > pH > aw.

Antimicrobial Activity of Pomegranate Peel Extracts as Affected by Cultivar.

BACKGROUND Some studies have reported that different parts of the pomegranate fruit, especially the peel, may act as potential antimicrobial agents and thus might be proposed as a safe natural alternative to synthetic antimicrobial agents. The high tannin content, especially punicalagin, found in pomegranate extracts, has been reported as the main compound responsible for such antimicrobial activity. Because the pomegranate peel chemical composition may vary with the type of cultivar (sweet, sour-sweet and sour), pomegranates may also differ with respect to their antimicrobial capacity. RESULTS The extract from PTO8 pomegranate cultivar peel had the highest antimicrobial activity, as well as the highest punicalagins (α and β) and ellagic acid concentrations. In the results obtained from both antibacterial and antifungal activity studies, the sour-sweet pomegranate cultivar PTO8 showed the best antimicrobial activity, and the highest ellagic acid concentrations. CONCLUSION The results of the present study suggest that ellagic acid content has a significant influence on the antimicrobial activity of the pomegranate extracts investigated. The pomegranate peel of the PTO8 cultivar is a good source of antifungal and antibacterial compounds, and may represent an alternative to antimicrobial agents of synthetic origin.

Chemical composition, cytotoxic and antioxidative activities of ethanolic extracts of propolis on HCT-116 cell line.

BACKGROUND Propolis is a complex resinous sticky substance that honeybees collect from buds and exudates of various plants. Owing to its versatile biological and pharmacological activities, propolis is widely used in medicines, cosmetics and foods. The aim of this study was to evaluate the cytotoxic and antioxidative effects of various ethanolic extracts of propolis (EEPs) on human colon cancer cell line HCT-116 and compare them with their composition determined by HPLC-DAD. RESULTS The most abundant flavonoids in all samples were chrysin, pinocembrin and galangin (12.697-40.811 µg mg⁻¹), while the main phenolic acids were caffeic acid, ferulic acid and isoferulic acid. Dose- and time-dependent inhibition of growth of HCT-116 cells was observed for all propolis samples, with IC₅₀ values ranging from 26.33 to 143.09 µg mL⁻¹. Differences in cytotoxic activity of propolis samples were associated with differences in their composition. All EEP samples reduced both superoxide anion radical and nitrite levels and also had strong DPPH-scavenging activity. CONCLUSION All tested propolis samples had pronounced cytotoxic and antioxidative activities.

Surface adhesins and exopolymers of selected foodborne pathogens

The ability of bacteria to bind different compounds and to adhere to biotic and abiotic surfaces provides them with a range of advantages, such as colonization of various tissues, internalization, avoidance of an immune response, and survival and persistence in the environment. A variety of bacterial surface structures are involved in this process and these promote bacterial adhesion in a more or less specific manner. In this review, we will focus on those surface adhesins and exopolymers in selected foodborne pathogens that are involved mainly in primary adhesion. Their role in biofilm development will also be considered when appropriate. Both the clinical impact and the implications for food safety of such adhesion will be discussed.

Antibacterial activity against Clostridium genus and antiradical activity of the essential oils from different origin.

In the present study, the antimicrobial and antiradical activities of 15 essential oils were investigated. The antimicrobial activities were determined by using agar disc diffusion and broth microdilution methods against Clostridium genus and antioxidant properties of essential oils by testing their scavenging effect on DPPH radicals activities. We determined the antibacterial activity of Clostridium butyricum, Clostridium hystoliticum, Clostridium intestinale, Clostridium perfringens and Clostridium ramosum. We obtained the original commercial essential oils samples of Lavandula angustifolia, Carum carvi, Pinus montana, Mentha piperita, Foeniculum vulgare Mill., Pinus sylvestris, Satureia montana, Origanum vulgare L. (2 samples), Pimpinella anisum, Rosmarinus officinalis L., Salvia officinalis L., Abies alba Mill., Chamomilla recutita L. Rausch and Thymus vulgaris L. produced in Slovakia (Calendula a.s., Nova Lubovna, Slovakia). The results of the disk diffusion method showed very high essential oils activity against all tested strains of microorganisms. The best antimicrobial activity against C. butyricum was found at Pimpinella anisum, against C. hystoliticum was found at Pinus sylvestris, against C. intestinale was found at Satureia hortensis L., against C. perfringens was found at Origanum vulgare L. and against C. ramosum was found at Pinus sylvestris. The results of broth microdilution assay showed that none of the essential oils was active against C. hystoliticum. The best antimicrobial activity against C. butyricum was found at Abies alba Mill., against C. intestinale was found at Abies alba Mill., against C. perfringens was found at Satureia montana and against C. ramosum was found at Abius alba and Carum carvi. Antioxidant DPPH radical scavenging activity was determined at several solutions of oil samples (50 μL.mL−1–0.39 μL.mL−1) and the best scavenging effect for the highest concentration (50 μL.mL−1) was observed. The antioxidant properties were different in particular plant species. The highest% of inhibition after 30 min. of reaction was observed at Origanum vulgare (93%), Satureia montana (90.66%) and Lavandula augustifolia (90.22%).

Heavy Metals Content and Microbiological Quality of Carp (Cyprinus carpio, L.) Muscle from Two Southwestern Slovak Fish Farms

The purpose of this study was to assess concentration and correlation of selected heavy metals and level of some microbiological indicators (total bacteria count-TBC, mesophilic anaerobic sporulating bacteria-MASB) in the muscle of common carp (Cyprinus carpio). Fish were collected by seine net within the frame of pondfishing, in October (pond Horné Obdokovce--pond A) and December (pond Budmerice--pond B) 2003. Concentrations of selected metals were measured using an atomic absorption spectrophotometer Pye Unicam SP9. The concentrations of metals (mg/kg wet weight basis) ranged as follows: Fe 3.47-15.15; Mn 0.14-0.42; Zn 3.47-9.52; Cu 0.24-1.32; Co 0.05-0.17; Ni 0.07-0.42; Cr 0.08-0.19; Pb 0.11-0.30; and Cd 0.01-0.05. The count of MASB and TCB (in CFU/g) varied as follow: 1.12-7.76 x 10(3) and 0.03-7.59 x 10(6), respectively. Significant differences (P < 0.05) for Cu, Ni, and Cr bioaccumulation, as well as for count of MASB between ponds were recorded. An opposite trend of bioaccumulation was found in only 12 of 36 cases. Positive correlations between counts of MASB and accumulated heavy metals, except Ni and negative correlations between TBC and accumulated heavy metals, except Cr, were recorded, (P < 0.05) for TBC-Mn and TBC-Cd relationship. Lead concentrations exceeded the maximum values allowed concentration in Slovakia by Codex Alimentarius (0.2 mg/kg) by 60% for pond-A and 40% for pond-B. The level of MASB count exceeded the maximum allowed concentration in 100% by both ponds. On average, the order of metal concentrations in the fish muscle was: (Pond-A) Fe > Zn > Cu > Mn > Pb > Cr > Ni > Co > Cd; (Pond-B) Fe > Zn > Cu > Mn > Pb > Ni > Cr > Co > Cd.