Antagonistički potencijal i sposobnost stvaranja biofilma bakterija Enterococcus spp. izoliranih iz sira u Srbiji

In this paper, the presence, biochemical and physiological characteristics of the members of genus Enterococcus, isolated from traditionally made Serbian cheese, were investigated. The members were identified as: Enterococcus faecium (8 isolates), E. faecalis (14 isolates), E. hirae (4 isolates) and E. durans (4 isolates), using the biochemical tests and MALDI-TOF mass spectrometry. All the tested isolates showed good acidification ability in pure and enriched milk. The antagonism of enterococci on the growth of Escherichia coli ATCC 25922, Proteus mirabilis ATCC 12453, Klebsiella oxytoca KGPMF1, Klebsiella ornithinolytica KGPMF8 and Aeromonas hydrophila, as well as their ability to form biofilms, were examined. The tested isolates showed moderate inhibitory activity (10-22 mm) on the growth of the indicator strains. Among all the isolates, only E. hirae KGPMF9 and E. faecium KGPMF14 showed the ability of biofilm formation. The results provide a basis for further research into the possible practical application of the isolated enterococci.U ovom je radu istražena prisutnost te biokemijske i fiziološke značajke bakterija iz roda Enterococcus, izoliranih iz domaćeg sira u Srbiji, proizvedenoga na tradicionalan način. Biokemijskim testom i masenom spektrometrijom MALDI-TOF dokazani su Enterococcus faecium (8 izolata), E. faecalis (14 izolata), E. hirae (4 izolata) i E. durans (4 izolata). Svi su analizirani izolati pokazali dobru sposobnost acidifikacije u sirovu i obogaćenom mlijeku. Istraženi su antagonizam enterokoka na rast bakterija Escherichia coli ATCC 25922, Proteus mirabilis ATCC 12453, Klebsiella oxytoca KGPMF1, Klebsiella ornithinolytica KGPMF8 i Aeromonas hydrophila te njihova sposobnost stvaranja biofilma. Analizirani izolati pokazali su umjerenu inhibicijsku aktivnost (10 - 22 mm) u rastu navedenih sojeva. Među izolatima samo su E. hirae KGPMF9 i E. faecium KGPMF14 pokazali sposobnost stvaranja biofilma. Rezultati ove studije daju temelj za daljnja istraživanja mogućnosti praktične primjene izoliranih enterokoka

In vitro procjena antimikrobnog potencijala bakterije Streptococcus uberis izolirane iz autohtonog sira jugoistočne Srbije

Streptococcus uberis is an environmental bacterium responsible for bovine mastitis. It is occasionally described as a human pathogen. In our study, the isolation was undertaken of lactic acid bacteria from a local cheese from Southeastern Serbia, produced in a traditional way. S. uberis (7 isolates) and S. agalactiae (1 isolate) were isolated from the cheese samples taken in the summer. The biochemical and physiological characteristics of the isolates were examined. Using tetracycline, chloramphenicol, novobiocin and rifampicin, the antibiotic susceptibility of the isolates was evaluated. The results demonstrated that all the isolates were susceptible to all the tested antibiotics, with a growth inhibition zone from 36-48 mm. Also, the antagonism was examined of S. uberis KGPMF1-7 and S. agalactiae KFPMF8 isolates on the growth of Escherichia coli ATCC 25922, Proteus mirabilis ATCC 12453, Klebsiella oxytoca KGPMF1, Klebsiella ornithinolytica KGPMF8 and Aeromonas hydrophila, as indicator stains. The results were compared with the activities of chloramphenicol, streptomycin and tetracycline on the tested indicator stains. The strongest antagonism was demonstrated by all Streptococcus isolates on the growth of K. oxytoca KGPMF1 (growth inhibition zone from 12-20 mm) and the A. hydrophila (growth inhibition zone from 13-20 mm). When these results were compared with the results of the sensitivity of tested indicator stains to antibiotics, S. uberis KGPMF1-7 and S. agalactiae KGPMF8 isolates showed a moderate antagonistic effect. Due to the specific way cheese is made in from Sokobanja, these isolates probably originate from cows’ udders.Streptococcus uberis jest okolišna bakterija odgovorna za mastitis kod goveda, a povremeno se opisuje kao patogen u ljudi. U ovom su istraživanju izolirane bakterije mliječne kiseline iz autohtonog sira jugoistočne Srbije, proizvedenog na tradicionalni način. S. uberis (7 izolata) i S. agalactiae (1 izolat) izolirani su iz uzoraka sira prikupljenih tijekom ljeta. Analizirane su biokemijske i fiziološke značajke izolata. Koristeći se tetraciklinom, kloramfenikolom, novobiocinom i rifampicinom, procijenjena je osjetljivost izolata na antibiotike. Rezultati su pokazali da su svi izolati osjetljivi na sve istražene antibiotike, sa zonom inhibicije rasta od 36 do 48 mm. Također, istražen je antagonizam S. uberis KGPMF1-7 i S. agalactiae KFPMF8 izolata na rast Escherichia coli ATCC 25922, Proteus mirabilis ATCC 12453, Klebsiella oxytoca KGPMF1, Klebsiella ornithinolytica KGPMF8 i Aeromonas hydrophila kao indikatorskih sojeva. Rezultati su uspoređeni s aktivnostima kloramfenikola, streptomicina i tetraciklina na testiranim indikatorskim sojevima. Najjači antagonizam pokazali su svi Streptococcus izolati na rast K. oxytoca KGPMF1 (zona inhibicije rasta od 12 do 20 mm) i A. hydrophila (zona inhibicije rasta od 13 do 20 mm). Usporedbom tih rezultata s rezultatima osjetljivosti istraženih indikatorskih sojeva na antibiotike, za S. uberis KGPMF1-7 i S. agalactiae KGPMF8 izolate utvrđen je umjeren antagonistički učinak. Ti izolati vjerojatno potječu iz vimena krava i u siru iz Sokobanje prisutni su zbog specifičnosti njegove pripreme

Biological activities of extracts from cultivated Granadilla Passiflora alata

Research conducted in this study showed the influence of ethanol, acetone and ethyl acetate extracts of the outgrowth of cultivated Passiflora alata on microorganisms, as well as the antioxidant activity and the concentrations of total phenols, flavonoids and tannins. In vitro antimicrobial activities of extracts were studied on 27 species of microorganisms, of which 17 species of bacteria and 10 species of fungi. The strongest antimicrobial activity was detected on G+ bacteria while the activities on other species were moderate. Ethyl acetate extract showed the strongest effect. The concentrations of total phenols were examined by using Folin-Ciocalteu reagent and the obtained values ranged from 14.04 to 34.22 mg GA/g. By using aluminium chloride method, the concentrations of flavonoids were obtained and the values ranged from 33.19 to 62.30 mg RU/g. In determining the amount of tannins we used the method with buthanol-HCl reagent and the obtained value was 5.1 % of dry matter. The efficiency of antioxidation, which we identified through the reduction of DPPH, was in the range from 808.69 to 1107.79 μg/ml for a particular IC50, and AAI values were between 0.07 and 0.10. The best parameters were shown by ethanol extract. All data were statistically analyzed. Overall, extracts showed potential for further investigation and use

Immortelle (Xeranthemum annum L.) as a natural source of biologically active substances

Antioxidant and antimicrobial effects, total phenolic content and flavonoid concentrations of methanolic, acetone and ethyl acetate extracts from Xeranthemum annuum L. were investigated in this study. The total phenolic content was determined using Folin-Ciocalteu reagent and ranged between 101.33 to 159.48 mg GA/g. The concentration of flavonoids in various X. annuum extracts was determined using spectrophotometric method with aluminum chloride and the results varied from 22.25 to 62.42 mg RU/g. Antioxidant activity was monitored spectrophotometrically using DPPH reagent and expressed in terms of IC50 (μg/ml), and it ranged from 59.25 to 956.81 μg/ml. The highest phenolic content and capacity to neutralize DPPH radicals were found in the acetone extract. In vitro antimicrobial activity was determined by microdilution method. Minimum inhibitory concentration (MIC) and minimum microbicidal concentration (MMC) have been determined. Testing was conducted against 24 microorganisms, including 15 strains of bacteria (standard and clinical strains) and 9 species of fungi. Statistically significant difference in activity between the extracts of X. annuum L. was observed and the acetone extract was found most active. The activity of acetone extract was in accordance with total phenol content and flavonoid concentration measured in this extract. The tested extracts showed significant antibacterial activity against G+ bacteria and weak to moderate activity against other microorganisms. Based on the obtained results, X. annuum can be considered as a rich natural source of polyphenolic compounds with very good antioxidant and antimicrobial activity

The influence of environmental factors on the planktonic growth and biofilm formation of Escherichia coli

In this study, the effects of environmental factors (different media, temperature, pH, salt and sugar concentrations) on the planktonic growth, biofilm formation and formed biofilm of Escherichia coli KGPMF 16 and Escherichia coli KGPMF 17 were investigated. Tested bacteria were isolated from traditionally made cheese produced in Southeastern Serbia (Sokobanja region). The influence on planktonic growth, biofilm formation and formed biofilm was determined using spectrophotometric method. The limiting factors for the planktonic growth and biofilm formation were temperature of 4 °C and all tested concentrations of salt. The growth of tested bacteria was higher in media enriched with lactose than in media containing glucose. TSB was more congenial media to the planktonic growth of bacteria than MHB broth. None of the tested bacteria demonstrated the ability to form biofilm at 4 °C and 44 °C. Only E. coli KGPMF 17 showed ability to form biofilm in TSB at 37 °C. Different concentrations of salt, glucose and lactose exhibited inhibitory effect on biofilm formation, but all tested concentrations of lactose showed stimulating effect on formed biofilm of E. coli KGPMF 17. These results contribute to better understanding of the effects of environmental factors on the development of E. coli in cheese

Heavy metal tolerance and removal efficiency of the Rhodotorula mucilaginosa and Saccharomyces boulardii planktonic cells and biofilm

The impact of heavy metals, cadmium (Cd2+), zinc (Zn2+) and nickel (Ni2+) on planktonic cells and biofilm of Rhodotorula mucilaginosa and Saccharomyces boulardii was examined. The metal tolerance testing was performed by MBECTM-HTP assay. The minimum inhibitory concentration (MICp) and minimum lethal concentration (MLCp) were determined as well as the minimum biofilm eradication concentration (MBEC). Biofilm was more tolerant on the presence of heavy metals than the planktonic cells. The planktonic cells of R. mucilaginosa were tolerant to high concentrations of Cd2+, Zn2+ and Ni2+, while the planktonic cells of S. boulardii tolerated Zn2+, exclusively. The R. mucilaginosa biofilm was tolerant to all of the tested metal concentrations and the obtained results were confirmed by fluorescence microscopy. S. boulardii did not show ability of biofilm formation. Metal removal efficiency of the R. mucilaginosa planktonic cells and biofilm were also tested. The R. mucilaginosa biofilm showed higher efficiency in metals removing compared to the planktonic cells. Until now, the heavy metal tolerance and the removal efficiency (Cd2+, Zn2+ and Ni2+) analyzes were performed solely on planktonic cells of Rhodotorula species. In this study, we investigated the metal removal efficiency of R. mucilaginosa planktonic cells and biofilm and compared the obtained results

Great horsetail (Equisetum telmateia Ehrh.)

This paper deals with the antioxidant and antimicrobial activity, total phenolic content and concentrations of flavonoids of Equisetum telmateia extracts. Total phenolic content was determined with Folin-Ciocalteu reagent and it ranged between 129.0 to 262.7 mg GA/g. The concentration of flavonoids in various extracts of E. telmateia was determined using spectrophotometric method with aluminum chloride and obtained results varied from 112.6 to 199.8 mg RU/g. Antioxidant activity was monitored spectrophotometrically and expressed in terms of IC50 (μg/ml), and its values ranged from 33.4 to 982.2 μg/ml. The highest phenolic content, concentrations of flavonoids and capacity to neutralize DPPH radicals were found in the acetone extract. In vitro antimicrobial activity was determined using microdilution method. Minimum inhibitory concentration (MIC) and minimum microbicidal concentration (MMC) were also determined. Testing was performed on 22 microorganisms, including 15 strains of bacteria (standard and clinical strains) and 7 species of fungi. There were statistically significant differences in activity between the extracts of E. telmateia. Different effects were noticed against the bacteria and the methanol extract appeared to be most efficient. All the extracts showed significant antibacterial activity against G+ bacteria and weak to moderate activity against other microorganisms

Phytomedical investigation of Najas minor All. in the view of the chemical constituents

Plants are an abundant natural source of effective antibiotic compounds. Phytomedical investigations of certain plants haven’t still been conducted. One of them is Najas minor (N. minor), an aquatic plant with confirmed allelopathy. Research conducted in this study showed the influence of water and ethyl acetate extracts of N. minor on microorganisms, in the view of chemical profiling of volatile constituents and the concentrations of total phenols, flavonoids and tannins. Antimicrobial activity was defined by determining minimum inhibitory and minimum microbicidal concentrations using microdilution method. Influence on bacterial biofilm formation was performed by tissue culture plate method. The total phenolics, flavonoids and condensed tannins were determined by Folin-Ciocalteu, aluminum chloride and butanol-HCl colorimetric methods. Chemical profiling of volatile constituents was investigated by GC and GC-MS. Water extract didn't have antimicrobial activity below 5000 µg/mL. Ethyl acetate extract has shown strong antimicrobial activity on G+ bacteria - Staphylococcus aureus PMFKGB12 and Bacillus subtilis (MIC < 78.13 µg/mL). The best antibiofilm activity was obtained on Escherichia coli ATCC25922 (BIC50 at 719 µg/mL). Water extract had higher yield. Ethyl acetate extract had a significantly greater amount of total phenolics, flavonoids and tannins. As major constituent hexahydrofarnesyl acetone was identified. The ethyl acetate extract effected only G+ bacteria, but the biofilm formation of G-bacteria was suppressed. There was a connection between those in vivo and in vitro effects against pathogenic bacterial biofilm formation. All of this points to a so far unexplored potential of N. minor

Microbiological index of water quality (mWQI) tested on the Gruza reservoir

The present paper considers formulation of a criterion for estimation of water quality from microbiological parameters (mWQI) based on comparison with WQI using certain elements of fuzzy logic on the example of the Gruza Reservoir. Quantitative estimation of water quality from five important microbiological parameters, the microbiological water quality index (mWQI) is realized using the proposed simple procedure. In view of the fact that the bulk of analyzed water samples belonged to class II, in future investigations the present method will be tested on lakes and reservoirs with a different degree of quality dispersion

Melilotus albus and Dorycnium herbaceum extracts as source of phenolic compounds and their antimicrobial, antibiofilm, and antioxidant potentials

Melilotus albus Medic. and Dorycnium herbaceum Vill. (Fabaceae) acetone, ethyl acetate, and ethanol extracts were investigated for their in vitro antimicrobial, antibiofilm, and antioxidant activity with quantification of phenolic compound contents. In general, D. herbaceum extracts showed better antibacterial and antioxidant activity than M. albus extracts. Bacteria Bacillus subtilis, Staphylococcus aureus ATCC 25923, Pseudomonas aeruginosa, and Proteus mirabilis were the most susceptible with the minimum inhibitory concentrations (MICs), determined by microdilution method, between 1.25–10 mg/mL. Antifungal activity was lower with the detectable MICs at 10 mg/mL and 20 mg/mL. The plant extracts, using the crystal violet assay, inhibit P. aeruginosa biofilm formation in concentration range from 5 mg/mL to 20 mg/mL whereas the effect on mature bacterial biofilm was lower. The antioxidant activity was evaluated using 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radicals scavenging and reducing power model systems. The intensity of DPPH radicals scavenging activity, expressed as half maximal effective concentration (EC50) values, was from 84.33 μg/mL to >1000 μg/mL. The extracts demonstrated reduced power in a concentration-dependent manner, with ethanol extract as the most active. The total phenols, flavonoids, and proanthocyanidins were determined spectrophotometrically while total extractable tannins were obtained by precipitation method. The phenolic compounds showed differences in their total contents depending on solvents polarities and plant species. Although the plants M. albus and D. herbaceum have not yet been fully explored, these results contribute better understanding of their biotic properties and potential application as antimicrobial and antioxidant agents