Antimicrobial activity of actinomycetes and characterization of actinomycin-producing strain KRG-1 isolated from Karoo, South Africa

In the present study we reported the antimicrobial activity of actinomycetes isolated from aridic soil sample collected in Karoo, South Africa. Eighty-six actinomycete strains were isolated and purified, out of them thirty-four morphologically different strains were tested for antimicrobial activity. Among 35 isolates, 10 (28.57%)showed both antibacterial and antifungal activity. The ethyl acetate extract of strain KRG-1 showed the strongest antimicrobial activity and therefore was selected for further investigation. The almost complete nucleotide sequence of the 16S rRNA gene as well as distinctive matrix-assisted laser desorption/ionization-time-of-flight/mass spectrometry (MALDI-TOF/MS) profile of whole-cell proteins acquired for strain KRG-1 led to the identification of Streptomyces antibioticus KRG-1 (GenBank accession number: KX827270). The ethyl acetate extract of KRG-1 was fractionated by HPLC method against the most suppressed bacterium Staphylococcus aureus (Newman). LC//MS analysis led to the identification of the active peak that exhibited UV-VIS maxima at 442 nm and the ESI-HRMS spectrum showing the prominent ion clusters for [M-H2O+H]+ at m/z 635.3109 and for [M+Na]+ at m/z 1269.6148. This information could be assigned to chromopeptide lactone antibiotic - actinomycin. Our results suggest that unexplored soils could be an interesting source for exploring antibacterial secondary metabolites

Effects of Drought on Microbial Activity in Rhizosphere, Soil Hydrophobicity and Leaching of Mineral Nitrogen from Arable Soil Depending on Method of Fertilization

This work presents the first results from the long-term laboratory experiment dealing with impact of drought on soil properties. Three groups of the treatment (A, B and C) with different regime of irrigation were prepared. The soil water content was maintained at 70 % of soil water holding capacity in group A, at 40 % in group B. In group C, soil water regime was maintained in the range of wilting point. Each group of the experiment was divided into three variants (A1 = B1, C1; A2 = B2, C2 etc.) with three repetitions: Variants A1 (B1, C1) were controls without addition of another fertilizer. Variants A2 (B2, C2) were fertilized with mineral nitrogen fertilizer DAM 390 (0.140 Mg of N per ha) and variants A3 (B3, C3) contained 45 g of Cp per a pot. The significant differences (ANOVA, P<0.05) in the leaching of mineral nitrogen and values of saturated hydraulic conductivity (Ksat) were found. The highest values of Ksat were found in variants (within each group) with addition of compost (A3, B3, C3). Conversely, the lowest values of Ksat were found in variants with addition of mineral nitrogen. Low values of Ksat indicate an increased level of hydrophobicity in individual groups of the experiment. Moreover, all variants with compost addition showed lower amount of mineral nitrogen leaching and high level of microbial activity than variants without. This decrease of mineral nitrogen leaching was about 200 % in comparison with the control variant and about 300 % with variant, where mineral nitrogen was added. Based on these results, we can conclude that changes of soil water content directly have impact on microbial activity, soil hydrophobicity and loss of mineral nitrogen from the soil

Isolation, antimicrobial activity of myxobacterial crude extracts and identification of the most potent strains

Broad spectrum antimicrobial agents are urgently needed to fight frequently occurring multidrug-resistant pathogens. Myxobacteria have been regarded as “microbe factories” for active secondary metabolites, and therefore, this study was performed to isolate two bacteriolytic genera of myxobacteria, Myxococcus sp. and Corallococcus sp., from 10 soil/sand samples using two conventional methods followed by purification with the aim of determining the antimicrobial activity of methanol extracts against 11 test microorganisms (four Gram-positive, four Gram-negative, two yeasts and one fungus). Out of thirty-nine directly observed strains, 23 were purified and analyzed for antimicrobial activities. Based on the broth microdilution method, a total of 19 crude extracts showed antimicrobial activity. The range of inhibited wells was more important in the case of anti-Gram-positive-bacterial activity in comparison with the anti-Gram-negative-bacterial and antifungal activity. In light of the established degree and range of antimicrobial activity, two of the most active isolates (BNEM1 and SFEC2) were selected for further characterization. Morphological parameters and a sequence similarity search by BLAST revealed that they showed 99% sequence similarity to Myxococcus xanthus − BNEM1 (accession no. KX669224) and Corallococcus coralloides - SFEC2 (accession no. KX669225). As these isolates had antimicrobial activity, they could be considered for use in the development of antibiotics for pharmaceutical use

Effect of fertilization on biological activity of community of soil streptomycetes

The search for new natural mechanisms to inhibit the growth of phytopathogenic microorganisms has become widely widespread. Therefore, the main objective of the present study was determination of antimicrobial activities of actinomycetes isolated from agricultural soil, which was fertilized mainly by organic fertilizers, against 8 selected phytopathogenic strains. Among the actinomycetes, Streptomyces species have been extensively studied, because they have been recognized as an important source of secondary metabolites, which can suppress the growth of undesirable pests in crops. The results indicated that the richest source of Streptomyces colonies was soil fertilized with compost (103 x104 CFU*g-1 dry soil). On the basis of morphological signs, total of 65 isolates were selected and examined for antimicrobial activities. Isolates exhibited the best activity against Gram negative bacterium Clavibacter michiganensis subsp. sepedonicus, disease agent of “ring rot” of potatoes and against fungus Fusarium poae, disease agent of Fusarium head blight of wheat. Twelve isolates exhibited promising broad-spectrum activity against tested organisms. On the basis of results, six of them were selected for further screening. Comparison of polyphasic studies with available literature led to identification of biological active strains S. olivochromogenes (13SC11), S. avermitilis (13SC2), S. rishiriensis (13SC13), S. globisporus (13SC19), S. sampsonii (13SPC10) and S. avidinii (13SPC4). After quantification analysis of various enzymes, tested isolates produced alkaline phosphatase, leucinearylamidase, valinearylamidase, acid phosphatase, naphtol-AS-BI-phosphohydrolase, glucosidase in high values (>40 nmol) and were positive for nitrate reduction, hydrolysis of gelatin, urease, and esculin. These isolates can be used in the development of new biopesticides anf biofertilizers with antibacterial and antifungal effect.Hľadanie nových prirodzených mechanizmov na potlačenie rastu fytopatogénnych mikroorganizmov sa stalo veľmi rozšíreným. Z tohto dôvodu je hlavným cieľom tejto štúdie detekcia antimikrobiálnej aktivity aktinomycét izolovaných z poľnohospodársky využívanej pôdy hnojenej najmä organickými hnojivami, voči 8 vybraným fytopatogénnym mikroorganizmom. Spomedzi aktinomycét, sme sa v tejto štúdii zamerali na streptomycéty, pretože patria medzi významných producentov sekundárnych metabolitov, ktoré môžu potlačiť rast neželaných škodcov úrody. Výsledky ukázali, že najbohatším zdrojom kolónií streptomycét bola pôda hnojená kompostom (103 x104 KTJ*g-1 sušiny). Na základe zistených morfologických znakov sme na štúdium antimikrobiálnej aktivity vybrali 65 izolátov. Izoláty vykazovali najlepší účinok voči baktérii Clavibacter michiganensis subsp. sepedonicus, pôvodcu choroby “kružkovatosti” zamiakov a voči hube Fusarium poae, pôvodcu choroby fuzáriovej sneti pšenice. 12 izolátov vykazovalo širokospektrálne účinky voči testovaným organizmom, 6 z nich bolo vybraných na ďalšie analýzy. Porovnanie ich polyfázických znakov s dostupnou literatúrou viedlo k identifikácii produkčných druhov S. olivochromogenes (13SC11), S. avermitilis (13SC2), S. rishiriensis (13SC13), S. globisporus (13SC19), S. sampsonii (13SPC10) a S. avidinii (13SPC4). Tieto izoláty sa vyznačovali tiež vysokou produkciou alkalickej fosfatázy, leucínarylamidázy, valínarylamidázy, kyslej fosfatázy, naftol-AS-BI-fosfohydrolázy a glukozídazy (> 40 nmol) a boli pozitívne pri redukcii dusičnanov, hydrolýzy želatíny, produkcii ureázy a eskulínu. Všetky tieto izoláty môžu byť použité pri vývoji nových biopesticídov a biohnojív s antifungálnymi a antibakteriálnymi účinkami

Soil myxobacteria as a potential source of polyketide-peptide substances.

Myxobacteria, a group of antimicrobial producing bacteria, have been successfully cultured and characterized from ten soil samples collected from different parts of Slovakia. A total of 79 myxobacteria belonging to four genera (Myxococcus, Corallococcus, Sorangium, and Polyangium) were isolated based on aspects of their life cycle. Twenty-five of them were purified, fermented, and screened for antimicrobial activities against 11 test microorganisms. Results indicated that crude extracts showed more significant activities against Gram-positive than against Gram-negative bacteria or fungi. Based on a higher degree and broader range of antimicrobial production, the two most potential extracts (K9-5, V3-1) were selected for HPLC fractionation against Micrococcus luteus and Staphylococcus aureus and LC/MS analysis of potential antibiotic metabolites. The analysis resulted in the identification of polyketide-peptide antibiotics, namely corallopyronin A and B (K9-5) and myxalamid B and C (V3-1), which were responsible for important Gram-positive activity in the observed strains. A sequence similarity search through BLAST revealed that these strains showed the highest sequence similarity to Corallococcus coralloides (K9-5, NCBI accession number KX256198) and Myxococcus xanthus (V3-1, NCBI accession number KX256197). Although screening of myxobacteria is laborious, due to difficulties in isolating cultures, this research represented the first report covering the isolation and cultivation of this challenging bacterial group from Slovakian soils as well as the screening of their antimicrobial activity, cultural identification, and secondary metabolite identification

Effect of fertilization on biological activity of community of soil streptomycetes

The search for new natural mechanisms to inhibit the growth of phytopathogenic microorganisms has become widely widespread. Therefore, the main objective of the present study was determination of antimicrobial activities of actinomycetes isolated from agricultural soil, which was fertilized mainly by organic fertilizers, against 8 selected phytopathogenic strains. Among the actinomycetes, Streptomyces species have been extensively studied, because they have been recognized as an important source of secondary metabolites, which can suppress the growth of undesirable pests in crops. The results indicated that the richest source of Streptomyces colonies was soil fertilized with compost (103 x104 CFU*g-1 dry soil). On the basis of morphological signs, total of 65 isolates were selected and examined for antimicrobial activities. Isolates exhibited the best activity against Gram negative bacterium Clavibacter michiganensis subsp. sepedonicus, disease agent of “ring rot” of potatoes and against fungus Fusarium poae, disease agent of Fusarium head blight of wheat. Twelve isolates exhibited promising broad-spectrum activity against tested organisms. On the basis of results, six of them were selected for further screening. Comparison of polyphasic studies with available literature led to identification of biological active strains S. olivochromogenes (13SC11), S. avermitilis (13SC2), S. rishiriensis (13SC13), S. globisporus (13SC19), S. sampsonii (13SPC10) and S. avidinii (13SPC4). After quantification analysis of various enzymes, tested isolates produced alkaline phosphatase, leucinearylamidase, valinearylamidase, acid phosphatase, naphtol-AS-BI-phosphohydrolase, glucosidase in high values (>40 nmol) and were positive for nitrate reduction, hydrolysis of gelatin, urease, and esculin. These isolates can be used in the development of new biopesticides anf biofertilizers with antibacterial and antifungal effect

Vplyv vermikompostu na zmeny bakteriálneho spoločenstva v rizosfére kukurice

The aim of the study was to observe changes in the diversity of bacterial community in maize rhizosphere influenced by organic and mineral fertilization. Four variants of fertilization were tested - vermicompost (VC) at recommended annual dose 40t*ha-1, doubled annual dose of VC, recommended dose of ammonium saltpeter with dolomite (LAD 27) and combination of VC and LAD 27. Experiment was conducted with potted maize plants in controlled conditions of greenhouse during 74 days. Using PCR-DGGE method, we investigated differences in total bacteria community as well as in community of ammonia oxidizing bacteria. Based on occurrence of operative taxonomic units (OTU) we found differences in bacterial species spectra among fertilization variants. The highest Shannon´s biodiversity index was observed in variant with VC addition in dose 80 t*ha-1.The fertilizers effect on diversity of ammonia oxidizing bacteria was not significant however in each variant with vermicompost addition was the occurrence of new specific OTU observed. This OTU was identified as Nitrosospira sp. It was proven that some bacterial species introduced to soil with vermicompost addition can survive for at least 74 days and these bacteria can influence basic functions of soil microbiocenosis in nitrogen cycle.Cieľom štúdie bolo sledovať zmeny diverzity bakteriálneho spoločenstva v rizosfére kukurice vplyvom organického a minerálneho hnojenia. Boli testované štyri varianty hnojenia – vermikompost (VC) v odporúčanej ročnej dávke 40t*ha-1, dvojitá ročná dávka VC, odporúčaná dávka liadku amónneho s dolomitom (LAD 27) a kombinácia VC a LAD 27. Experiment bol realizovaný na nádobách s rastlinami kukurice v kontrolovaných podmienkach skleníka počas 74 dní. Využitím metódy PCR – DGGE sme skúmali rozdiely v celkovej komunite baktérií ako aj komunite nitrifikačných baktérií. Na základe prítomnosti operatívnych taxonomických jednotiek (OTU) sme zistili rozdiely v spektrách bakteriálnych druhov v závislosti od variantov hnojenia. Najvyšší Shannonov index biodiverzity bol zaznamenaný vo variante s prídavkom VC v dávke 80 t*ha-1. Vplyv hnojenia na diverzitu nitrifikačných baktérií nebol preukazný, avšak v každom variante s prídavkom vermikompostu bol zaznamenaný výskyt nového špecifického OTU. Tento OTU bol identifikovaný ako Nitrosospira sp. Dokázalo sa, že niektoré druhy baktérií dodané do pôdy prídavkom vermikompostu sú schopné prežívať po dobu najmenej 74 dní a môžu ovplyvňovať základné funkcie pôdnej mikrobiocenózy v cykle dusíka