ANTAGONISTIC PROPERTIES OF THE BLACK SEA STREPTOMYCETES ISOLATED FROM THE FOULING OF SHELL ROCK AND MUSSELS

SummaryThe rapid emergence of resistance by bacterial and fungal pathogens is a serious problem in the health care system, which causes the search for new promising producers of antimicrobial natural products in various ecological niches. Aim. To determine the antagonistic activity of streptomycetes isolated from the biological fouling of natural shell rock and mussels of the Odesa gulf of the Black Sea. Methods. The antagonistic activity of 19 and 14 strains of streptomycetes isolated from the fouling of shell rock and mussels of the Odesa gulf, respectively, were investigated. Streptomycetes were pre-cultivated on agar media Gause 1, Gause 2 and oat agar with sea salt (2%) at a temperature of 30 °C for 10 days. Antagonistic activity against 12 test cultures was determined by the block method. Results. All isolated marine streptomycetes are antagonists of at least one strain of the indicator microorganism. Antibiotic activity depended on the source of the streptomycetes isolation, culture medium and properties of specific strains of both producers and test cultures. The best activity of streptomycetes strains from shell rock was shown after cultivation on Gause 1 medium, and streptomycetes from mussels – after cultivation on Gause 2 medium. The zones of no growth of sensitive indicators ranged from 12,4±0,3 mm to 20,6±0,2 mm (under the influence of streptomycetes from shell rock) and from 12,4±0,2 mm to 39,7±0,2 mm (under the influence of streptomycetes from mussels). Streptomyces sp. Lim 2.2 (strain from a shell rock) inhibited the growth of 8 test cultures, and strains from mussels Streptomyces sp. Myt 4b and Myt 7ch – 10 test cultures. Indicator strains of gram-positive bacteria were the most sensitive to all streptomycetes, in particular, strain Staphylococcus aureus ATCC 25923 was most inhibited by metabolites of Streptomyces spp. Myt 12a and Myt 12b. Conclusions. Antagonistic activity of streptomycetes isolated from the Black Sea depended on the source of isolation, pre-cultivation medium and properties of both producer strains and indicator microorganisms. The greatest activity of streptomycete strains from shell rock and mussels was shown after preliminary cultivation, respectively, on Gause 1 and Gause 2 media against gram-positive bacteria. The best antibiotic potential was found in strains of Streptomyces sp. Lim 2.2, Lim 4, Lim 5.1 and Lim 7.2, isolated from the fouling of shell rock, and strains of Streptomyces sp. Myt 7b, Myt 7ch, Myt 12a and Myt 12b isolated from mussels.</jats:p

THE COMPOSITION OF CELLULAR FATTY ACIDS OF ACTINOBACTERIA FROM THE SURFACES OF BIOLOGICAL GROWTH OF THE ODESA GULF OF THE BLACK SEA

Aim. Determination of fatty acid composition of actinobacteria isolated from the surfaces of biological growth of the Odesa gulf of the Black Sea, and their identification. Methods. The 31 isolated strains of actinobacteria were grown in TSB at 28 ° C and 150 rpm for 72 hours. Fatty acid methyl esters of the studied strains were determined according to the MIS Operating Manual on a gas chromatograph Agilent 7890, identification was performed using the identification system of microorganisms MIDI Sherlock. Results. Using chromatographic analysis of fatty acids, it was found that of the 27 studied strains of actinobacteria were identified to the genus Streptomyces, and the 4 strains - to the genus Nocardiopsis. It was found that the fatty acid profiles of the studied actinobacteria of the genus Nocardiopsis were dominated by fatty acids: 15:0 ANTEISO, 16:0 ISO, 17:0 ANTEISO, 18:1 CIS 9, and the fatty acid profiles of bacteria of the genus Streptomyces - 14:0 ISO, 15:0 ANTEISO, 16:0 ISO, 17:0 ANTEISO. Conclusions. Actinobacteria the surfaces of biological growth of the Odesa gulf of the Black Sea belong to the genera Streptomyces and Nocardiopsis, and their fatty acid profiles are characterized by the dominance of isomers of branched saturated fatty acids.</jats:p

RESISTANCE TO HEAVY METALS OF ACTINOBACTERIA ISOLATED FROM THE BIOLOGICAL FOULING OF SHELLFISH AND MUSSELS OF THE ODESA GULF OF THE BLACK SEA

SummaryEnvironmental pollution with heavy metals/metalloids is one of the most important environmental problems, which leads to the development of bioremediation strategies and the search for biomarkers to assess its condition. Aim. To determine the sensitivity to heavy metals of actinobacteria isolated from the biological fouling of natural shell rock and mussels of the Odesa gulf of the Black Sea. Methods. Thirty-four strains of actinobacteria isolated from fouling of shell rock and mussels of the Odesa gulf were used in the investigation. The sensitivity of the studied bacteria to heavy metal cations was determined on starch-casein agar by the disk diffusion method. Discs impregnated with salts solutions of Cu2+, Co2+, Ni2+, Cd2+, Zn2+ in concentrations of 0.001 mol/l, 0.01 mol/l, 0.05 mol/l, 0.1 mol/l, 0.5 mol/l and 1.0 mol/l were used. Results. The investigated actinobacteria showed variable sensitivity to heavy metals, which depended on the source of isolation, strain, type of metal and its concentration. All tested bacteria were most sensitive to Cd2+ (MIC was 0.001 mol/l), the most resistant to Zn2+ (MIC was higher than 1.0 mol/l for the vast majority of bacteria). At concentrations lower than the MIC, zinc stimulated the formation of aerial mycelium of almost all strains, and pigment formation increased in some of them. The sensitivity to heavy metals of actinobacteria isolated from shellfish decreased in the following sequence: Cd2+&gt;Cu2+&gt;Co2+&gt;Ni2+&gt;Zn2+, and in actinobacteria isolated from mussels – Cd2+&gt;Cu2+&gt;Ni2+&gt;Co2+&gt;Zn2+. Conclusions. Actinobacteria isolated from mussels are more sensitive to cadmium, cuprum, cobalt, nickel and zinc, compared with actinobacteria from shell rock. All studied strains were highly sensitive to Cd2+ (MIC Cd2+ for almost all strains was 0.001 mol/l) and resistant to Zn2+ in the concentration range of 0.001 mol/l – 0.5 mol/l.</jats:p

BIOSURFACTANTS SYNTHESIS BY PSEUDOMONAS AERUGINOSA BACTERIA ISOLATED FROM THE SURFACE OF MUSSELS OF THE BLACK SEA

Aim. Establishing of the ability to synthesize surface-active compounds by Pseudomonas aeruginosa bacteria isolated from the surface of Black Sea mussels. Methods. During the research several marine Pseudomonas spp strains isolated from petroleum hydrocarbon contaminated areas of Black Sea wereused: P. aeruginosa M1, P. aeruginosa M4 and P. aeruginosa PA01 as reference strain in suspension and biofilm cultures (LB and Giss media). Cultivation of Pseudomonas aeruginosa strains was performed at 37 °C for 120 and 168 hours. Planktonic culture growth was determined spectrophotometrically on the wave length 600 nm. Biofilm mass was determined spectrophotometrically on the wave length 592 nm by CV-test. The presence of surface-active compounds was determined in a drop test. The quantitative content of rhamnolipids was evaluated by the color reaction of rhamnose with orcin. Results. P. aeruginosa strains M1 and M4 isolated from Black Sea mussel’s surfaces synthesize 25% and 66% more surfactants than the reference strain PA01. All strains in Giss medium synthesized 10–20 times less rhamnolipids than in LB medium. In biofilm cultures the same biosurfactant synthesis dependence on the composition of the nutrient medium is observed as in suspension cultures. According to the intensity of rhamnolipid production in biofilm cultures, the studied strains can be arranged in the following row: P. aeruginosa M4 &gt; P. aeruginosa M1 &gt;&gt; P. aeruginosa PA01.Conclusions. The strains of P. aeruginosa isolated from the Black Sea are more efficient producers of rhamnolipids than the reference strain of P. aeruginosa PA01; the intensity of biosynthesis of surfactants significantly depends on the composition of the nutrient medium and the method of cultivation.</jats:p