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Soil microorganisms and their activities are essential for maintaining soil health and fertility. Microorganisms can be negatively affected by application of herbicides. Although effects of herbicides on microorganisms are widely studied, there is a lack of information for chloroacetamide herbicide dimethachlor. Thus, dimethachlor and well known linuron were applied to silty-loam luvisol and their effects on microorganisms were evaluated during112 days long laboratory assay. Dimethachlor and linuron were applied in doses 1.0 kg ha−1 and 0.8 kg ha−1 corresponding to 3.33 mg kg−1 and 2.66 mg kg−1 respectively. Also 100-fold doses were used for magnification of impacts. Linuron in 100-fold dose caused minor increase of respiration, temporal increase of soil microbial biomass, decrease of soil dehydrogenase activity, and altered microbial community. Dimethachlor in 100-fold dose significantly increased respiration; microbial biomass and decreased soil enzymatic activities. Microbial composition changed significantly, Proteobacteria abundance, particularly Pseudomonas and Achromobacter genera increased from 7 to 28th day. In-silico prediction of microbial gene expression by PICRUSt2 software revealed increased expression of genes related to xenobiotic degradation pathways. Evaluated characteristics of microbial community and activity were not affected by herbicides in recommended doses and the responsible use of both herbicides will not harm soil microbial community.

There is a growing interest in the issue of inoculation of rhizobacteria into the agricultural soil because this group of bacteria can increase productivity and quality of agriculturally important crop and contributes to the stability of agroecosystems. The aim of our work was to isolate and characterize of plant growth promoting traits (production of IAA, siderophores, phosphate solubilisation, antifungal activity) of rhizobacteria belonging to a group of plant growth promoting rhizobacteria (PGPR), and isolated from rhizosphere of maize (Zea mays L.) in luvisols. Quantitative representation of rhizobacteria of maize was 7.4 . 106 CFU.g-1 dry soil. A total of eleven species of maize rhizosphere where isolated and confirmed as PGPR in vitro. The all isolates showed positive indole-3-acetic acid (IAA) production ranging between 1.39 and 15.74 µg.ml-1. Seven strains (63.6 %) has been shown with low and 1 strain with intermediate solubilisation index of phosphates and the positive production of siderophores showed 7 isolates (63, 6 %). Except for the isolate KmiJP17B089, all others inhibited the growth of Sclerotinia sclerotiorum and Rhizoctonia solani by more than 50 %. In the case of Fusarium graminearum, on the other hand, we observed a very low inhibitory activity. The most active in observed traits were three isolates identified by the 16S rRNA gene sequencing and NCBI database using the BLAST resource as Bacillus altitudinis strain KmiJP17B089, Bacillus aryabhattai strain KmiJP17B090 and Bacillus megaterium strain KmiJP17B091. These results suggest the possibility to tested in vivo these Bacillus species as potential biological fertilizer to increase maize production.

Plant microbiomes are responsible for the growth, health, nutrition, and resistance to biotic and abiotic stress. Moreover, some plant microbiome members, especially the Enterobacteriaceae family, can act as human pathogens. In recent years, metagenomic analysis of amplified 16S rRNA gene sequenced on Illumina second generation sequencers became the most widely used method for bacterial microbiome studies. The 16S r RNA gene of the bacterium is phylogenetically similar to the 16S genes of chloroplasts and mitochondria since they have common prokaryotic ancestors. Thus, plant microbiome analysis is affected by unwanted non-target chloroplast and mitochondrial sequences. The solution to this contamination uses specially designed primer pairs that exclude chloroplasts and mitochondria and provide undistorted information. In this study, we analyzed chloroplast excluding primer 799R (reverse complement of 799F) which contains 4 mismatches of nucleotides against the chloroplast sequence modified for use with Illumina sequencers. We used primer 799R with universal bacterial forward primers 341F and 515F and compared the results obtained with 799R to the results obtained with the universal 806R primer. Microbiomes of 12 samples from 4 distinct parts of the tomato plants (Solanum lycopersicon L.) leaves, fruits, roots, and rhizosphere, were amplified using all combinations of these primers. The combination of universal forward primers 515F or 341F with the universal bacterial primer 806R amplified 60-85% of chloroplast sequences in samples of plant tissue. The 799R primer effectively reduced the number of chloroplast sequences to less than 1%. However, some sequences derived from mitochondria remained, with a higher proportion using the 341F primer (66-72%) than 515F (21-28%). The ability to describe microbial population diversity using 799R was similar to 806R, although there is a clear difference in the proportions of amplified microorganism groups using these primers. Analysis revealed the highest frequency of Enterobacteriaceae when primer pair 515F+799R was used. Based on the results, the combination of primers 515F+799R was the most appropriate, met all parameters, and can be recommended for routine analysis of plant microbiomes using second generation sequencers.

Inoculation of Streptomyces to improve oilseed rape (Brassica napus L.) yields and minimise the use of chemical fertilisers is a promising sustainable strategy. In this study, we isolated 72 actinobacterial strains from rhizosphere of oilseed rape and maize and from bulk soil for screening and characterising their antimicrobial activity. Nine promising strains, identified as Streptomyces sp. by morphology, physiological characteristics, and 16S rRNA gene sequencing, were selected for their plant growth-promoting traits and in planta experiments. The actinobacterial strains were positive for IAA production, siderophore production, and HCN production. In planta experiments were conducted by soaking the oilseed rape seeds in the actinobacterial suspension, followed by plant growth under controlled conditions in a cultivate chamber (22–28 °C, 8 h dark/16 h light, constant humidity 80%). We recorded root and shoot length (cm) and seedling fresh weight (g). For most of the abovementioned parameters, a significant enhancement was observed with strain KmiRC20A118 treatment. The length of the root increased by 53.14%, the shoot length increased by 65.6%, and the weight of the fresh plant by 60% compared to the control. The integrated application of PGPS (Plant Growth Promoting Streptomyces) from the rhizosphere of oilseed rape is a promising strategy to improve the growth of oilseed rape.

Plant diseases caused by pathogenic fungi are a severe problem for agriculture, and some organic fertilizers have shown a suppressive capacity due to the antagonistic action of microbial communities inhabiting those substrates. The purpose of this work was to isolate and identify actinomycetes from soil, compost and vermicompost able to antagonize phytopathogens. In total, out of 352 actinomycetes successfully recovered on Pochon medium and of which were selected representative 22 mycelium forming isolates, which were subjected to morphological and genotypic characterization. Genetic characterization based on 16S rDNA fragment sequencing revealed that, all the actinomycete isolates belong to the genus Streptomyces. The antifungal activity of isolates was tested against eight phytopathogenic fungi, and they were most activated against Rhizoctonia solani, Alternaria tenuissima, Aspergillus niger and Penicillium expansum. Isolates 51VK13 (Streptomyces sampsonii) and 12VK13 (Streptomyces flavovariabilis) isolated from soil amended with vermicompost showed the greatest antagonistic activity. Their bioactive potential was also confirmed by presence of genes for nonribosomal peptide synthetase (NRPS) and polyketide synthase types I and II (PKS-I and PKS-II). It can, therefore, be concluded that soil amended with organic fertilizers such as compost, and in particular vermicompost, provides a lower incidence of phytopathogens, especially fungi. Isolated actinomycetes have also shown high potential for the production of bioactive compounds useful in the control of plant diseases.

Species of the genus Beauveria are entomopathogenic fungi used in biological control to protect against insect pests. Identification of species is based on observation of morphological, phylogenetic and genetic differences. Currently, MALDI-TOF (Matrix Assisted Laser Desorption/Ionization- Time of Flight) mass spectrometry is often used in microbial diagnostic but identification of Beauveria species has not been tested before. The aim of this work was the possibilities of this method and to compare it with genetic analysis. Twenty strains of B. bassiana and B. pseudobassiana isolated from insect cadavers, mycoinsecticides and soil samples were used. Sequences of ITS (Internal Transcribed Spacer) region were used for genetic identification of strains. Sequences and results from protein analysis of isolates were compared with the reference strains of B. bassiana and B. pseudobassiana. Results of MALDI Biotyper softvare identification were correct for all strains. Identification was reliable with high log score (>2.1) for most of strains. Score value less than 1 was observed only in a single case - strain GHA isolated from mycoinsecticide Botanigard.The result of both analyses was also dendrogram (phylogenetic tree) which showed high degree of result consistence. MALDI-TOF identification of fungi from Beauveria genus is usable for fast and reliable identification of entomopathogenic fungi isolates acquired from environment and results can be utilized in conservation biocontrol strategies in agriculture and forest ecosystems.

Rapeseed is an important oil crop strongly dependent on high agrochemical inputs. Some pathogens, including Leptosphaeria maculans, cause blackleg disease and can drastically decrease yields. Microbial inoculants seem to be a promising solution to these problems. However, a selection of potent bacterial strains able to improve growth and/or suppress disease is needed. Endophytic bacteria (n = 38) isolated from rapeseed plants with exceptionally good growth were screened for plant growth promoting (PGP) traits and L. maculans antifungal activity. A majority of isolates (35) showed the ability to produce siderophores, 17 isolates solubilized phosphate, and 28 isolates inhibited the growth of L. maculans. The six most promising isolates belonging to Bacillus genera were characterized in detail and compared to two previously published PGP strains. Plant growth measured as total weight and root length of rapeseed seedlings was stimulated by all isolates in comparison to control. The best isolate, 1L6, preliminary identified as Bacillus pumilus showed the highest phosphate solubilization, IAA and HCN production, and growth promotion of plants. Isolates with high antifungal activity in screening showed good potential to suppress disease on plants, with 87% reduction of lesions caused by L. maculans. These strains are good candidates to be explored under field use either solely or in combination.

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.

The presented paper deals with the analysis of potential differences between organic waste compost (CBD), vermicompost (CVER) and mineral fertilizer (MF, 27% of N) applications affecting the quality of arable soil by influencing microbial activity therein. The selected types of compost represent alternatives to conventional organic fertilizers, which are, however, not available to Czech and Slovak farmers in sufficient amounts. Their mutual comparison and the comparison with organic fertilizers aim to provide farmers further information about their influence on arable land and thus to give them the possibility of deciding on the most suitable amendments. To demonstrate the effect of these amendments, six variants were prepared: one without the addition of fertilizers, two variants with the addition of 40 Mg/ha of CVER and CBD, one variant with the addition of double dosed CVER (80 Mg/ha), and the remaining two variants were fertilized only with MF (0.22 Mg/ha) and with the combination of CVER (0.20 Mg/ha) and MF (0.11 Mg/ha). Substrate induced respiration (SIR), basal respiration (BS), microbial carbon (Cmic) and enzymatic activities (hydrolysis of fluorescein diacetate&mdash, FDA, dehydrogenase activity&mdash, DHA, and phosphatase activity&mdash, PA) were used to evaluate the effect of CBD, CVER and MF application on the soil quality. Both organic and mineral amendments affected BS and SIR. The highest BS and SIR rates were found in variants with compost application (CVER and CBD). All variants treated with the mineral fertilizer showed the lowest level of enzyme activities, lower by about 30% in comparison with variants where CVER, CBD and the combination of MF and CVER were applied. We found insignificant differences between the individual types of compost. More importantly, we compared the situation at the beginning of the experiment and after its end. It was found that the application of mineral fertilizers automatically led to the deterioration of all enzymatic parameters, on average by more than 25%, as compared with the situation at the beginning of the experiment. However, when the mineral fertilizer dose was supplemented with organic amendments (CVER), this negative effect was eliminated or significantly reduced. Furthermore, both composts (CVER and CBD) positively affected plant biomass production, which reached a level of production enhanced by the MF. Results clearly showed that the application of both compost types could be used to improve soil quality in agriculture.

Bacteria with positive properties on plant vitality are also called PGPB (Plant Growth Promoting Bacteria). Their presence can be observed not only in the root area but also in the above-ground parts of plants like endophytic bacteria. The aim of our study was to characterize promoting features of bacteria from Bacillus genus and compare them with Pseudomonas simiae WCS417 (plant growth promoting strain). The work was carried out in locality Kolíňany near Nitra (40°26´46´´N, 79°58´56´´W) and root samples were taken from 6 randomly selected plants of maize (Zea mays L.) in vegetative plant growth stage BBCH 14-15. Bacteria isolated from plant roots were identified and tested to biochemical parameters. From the biochemical features, we observed the detection of siderophores, determination of indole-3-acetic acid (IAA), monitoring the ability to dissolve phosphates and antifungal activity. Bacterial suspensions were applied to maize seeds and tested in vivo controlled conditions. Tested isolates were identified as Bacillus flexus, Bacillus megaterium and Bacillus subtilis. All 3 strains achieved the middle – class of phosphate solubilization index (2.00 ≤ SI ˂ 4.00), produced phytohormone IAA and showed positive production of siderophores and inhibited growth of Fusarium culmorum to more than 50%. All differences between tested strains and control strain P. simiae WCS417 were also statistically confirmed. All strains showed positive results in monitoring plant growth promoting properties. The effect of three Bacillus strains on maize seeds in vivo conditions showed significant differences in root length (P

In laboratory assay, the diversity of bacteria and microscopic fungi and the community-level physiological profiling (CLPP) of microorganisms were observed after the addition of bio-sludge (40 t.ha -1 ) from a biogas station and addition of bio-fertlizers - AZOTER (10 dm -3 .ha -1 ) to the arable soil with PCR-DGGE and BIOLOG ® method (Eco Plates). The differences were recorded in the microbial diversity (bacteria and microscopic fungi) among variants according to the Shannon index. The differences in community of microscopic fungi were markedly higher among the soil samples with the additions of both bio-fertilizers compared to control soil samples. The occurrence of individual OTUs (operational taxonomic units) bacteria and microscopic fungi were different after 105 days of incubation from the status after the 1 st day of incubation. The community metabolic diversity (CMD) was influenced by the incubation time (105 days) as well, but not by application of bio-fertilizers. We observed a significant decrease (LSD test, P

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.

The aim of our work was to survey and state the representative values and range of the microbial biomass carbon (Cmic) and microbial respiration activities in arable soils of three types (Chernozem, Luvisol, Planosol) and in pasture grassland soil (Cambisol). In arable soils no significant differences between the soil types were found concerning the content of Cmic. Way of soil use significantly influenced Cmic with higher values on pasture grassland soil. In Cambisol, a relatively strong relationships between Cmic and Corg (r = 0.919) as well as between Cmic and Ntot (r = 0.922) were determined. The basal respiration (BR) was no affected neither by a soil type nor by a way of soil use. On all monitored sites microbes responded positively to the addition of glucose (potential respiration - PR), but the strongest response was observed on the soil types with good mineralisation ability, such as Chernozems and Luvisols. On pasture grassland soil, the substrate availability index (PR/BR) with values of 2.32 was observed, which is lower compared to 7.60 of arable soil. This fact to indicate that pasture grassland soil had more easy decomposable organic compounds than arable soils.